Queen Elizabeth II’s flag-draped coffin arrives in Scottish capital: ‘An extraordinary woman’

EDINBURGH, Scotland – Queen Elizabeth II’s flag-draped coffin has arrived at the Palace of Holyroodhouse, the British monarchy’s residence in the Scottish capital city of Edinburgh.

The arrival followed a slow, somber and regal procession through the Scottish countryside on Sunday. Mourners had packed city streets and lined rural roads to take part in a historic goodbye to the monarch who reigned for 70 years.

The hearse drove past piles of bouquets and other tributes as it led a seven-car cortege from her beloved summer estate Balmoral Castle, where the queen died Thursday at age 96, for a six-hour trip through Scottish towns to the palace in Edinburgh. The late queen’s coffin was draped in the Royal Standard for Scotland and topped with a wreath made of flowers from the estate, including sweet peas, one of the queen’s favorites.

Members of the Royal Regiment of Scotland carried the coffin past the queen’s only daughter, Princess Anne, and into the throne room. It will remain there until Monday afternoon so residence staff can pay their last respects.

“A sad and poignant moment as Her Majesty, The Queen leaves her beloved Balmoral for the final time,” the first minister of Scotland, Nicola Sturgeon tweeted. “Today, as she makes her journey to Edinburgh, Scotland will pay tribute to an extraordinary woman.”

King Charles III and his Queen Consort Camilla will travel to Edinburgh on Monday to take the coffin to St. Giles Cathedral on the city’s Royal Mile. The coffin will remain there for 24 hours before being flown to London on Tuesday. The coffin will be moved from Buckingham Palace on Wednesday to the Houses of Parliament to lie in state until a state funeral at Westminster Abbey on Sept. 19.

► Royal news right in your inbox: Subscribe to Keep Calm and Carry On, a newsletter answering all of your questions about the royal family following Queen Elizabeth’s death.

In the Scottish village of Ballater, where residents regard the royal family as neighbors, hundreds of people watched as the hearse it passed.

“She meant such a lot to people in this area. People were crying, it was amazing to see,” said Victoria Pacheco.

The procession was mostly met with muted scenes of respect. People stood in silence; some clapped politely, others pointed their phone cameras.

What’s next for the young royals? King Charles names Will and Kate the Prince and Princess of Wales

In Ballater, the Rev. David Barr said locals consider the royals as “neighbors” and try to treat them as locals when they spend summers in the Scottish Highlands.

When Elizabeth arrived at her summer estate, the royal part of her mostly stayed outside, he said. “She was able to be a wife, a loving wife, a loving mum, a loving gran and then later on a loving great-gran – and aunty – and be normal.”

Elizabeth Taylor of Aberdeen had tears in her eyes after the hearse passed through Ballater.

“It was very emotional. It was respectful,” she said. “She certainly gave service to this country, even up until a few days before her death.”

Protest breaks out in Edinburgh to ‘abolish the monarchy’

Just before the proclamation of King Charles III was read Sunday in Edinburgh, a protester appeared with a sign condemning imperialism and urging leaders to “abolish the monarchy.” She was taken away soon after by police. Reaction was mixed. One man shouted, “Let her go! It’s free speech!” while others yelled, “Have some respect!”

Still, there was also some booing in Edinburgh when Joseph Morrow, Lord Lyon King of Arms, finished his proclamation with the words “God save the king!”

Ann Hamilton, 48, said she thought it was “absolutely terrible” that people booed the royal family during the proclamation.

“There’s tens of thousands of people here today to show their respect. For them to be here, heckling through things, I think it was terrible. If they were so against it, they shouldn’t have come,” she said.

Queen Elizabeth’s death: Operation London Bridge and Operation Unicorn, explained

President Biden praises Queen Elizabeth II on anniversary of 9/11 

While marking the 21st anniversary of the Sept. 11, 2001, terrorist attacks, President Joe Biden paid tribute to the late Queen Elizabeth II for her support of the United States on that fateful day.

In a speech at the Pentagon, Biden cited the queen’s message to the U.S. shortly after the 9/11 attacks: “Grief is the price we pay for love.”

Earlier Sunday, Biden said he has not yet spoken to King Charles III since the death of his mother.

“I’ve spoken to him many times but I haven’t spoken to him since he’s become king,” Biden told reporters as he left Delaware to return to Washington for the 9/11 commemoration.

Biden is scheduled to travel to London, accompanied by first lady Jill Biden, to attend the queen’s funeral.

What kind of sovereign will King Charles III be? Not like his mother

Prince William promises to serve Wales with ‘humility and great respect’

Prince William, the new Prince of Wales, has told the First Minister of Wales, Mark Drakeford, he will serve the country “with humility and great respect.”

William was given the title when his father, King Charles III, the previous Prince of Wales, ascended to the throne following the death Thursday of Queen Elizabeth II.

William told Drakeford that he and his wife, Kate, have a “deep affection for Wales, having made their first family home in Anglesey, including during the earliest months of Prince George’s life.”

He said the couple would travel to Wales “very soon” and want “to do their part to support the aspirations of the Welsh people and to shine a spotlight on both the challenges and opportunities in front of them.”

The world reacts to Queen Elizabeth II’s death: King Charles III, Prince William, more pay tribute

King Charles III formally proclaimed new monarch in Scotland, Wales, Northern Ireland

King Charles III has been formally proclaimed the new monarch in Scotland, Wales and Northern Ireland.

The move came Sunday in the rest of the United Kingdom, a day after the same proclamation was made in London at a pomp-filled accession ceremony steeped in ancient tradition and political symbolism.

In Belfast, bells chimed and a bugler played before the proclamation was read. It was followed by a 21-gun salute and a military band playing the anthem, “God Save the King.” In Wales, a regimental mascot goat accompanied the 3rd Battalion of the Royal Welsh regiment at the ceremony at Cardiff Castle.

Earlier, proclamations were held in other parts of the Commonwealth – the group of former British Empire colonies – including Australia and New Zealand.

Even as he mourned Elizabeth, Charles was getting down to work. He met at Buckingham Palace with the secretary-general and other representatives of the Commonwealth, a group of nations that grapples with affection for the queen and lingering bitterness over their own colonial legacies, ranging from slavery to corporal punishment in African schools to looted artifacts held in British institutions.

Contributing: David Jackson, USA TODAY; The Associated Press

Unstoppable: The Extraordinary Life of Carmen Pacheco

Introduction:

Carmen Pacheco, an inspirational figure in the world of business and philanthropy, has led an extraordinary life filled with resilience, determination, and unwavering commitment. Born on February 15th, 1965, in a small town in Mexico, Carmen faced numerous challenges throughout her journey. However, she defied all odds and emerged as a prominent force in the corporate world. This biography aims to delve into the incredible story of Carmen Pacheco’s life, exploring her early years, professional achievements, and significant contributions to society.

Chapter 1: Early Years

Carmen Pacheco was raised in a modest household where she learned the value of hard work and perseverance from her parents. Her father worked as a farmer while her mother dedicated herself to taking care of their five children. Growing up with limited resources didn’t deter Carmen from dreaming big and aspiring for greatness.

From an early age, Carmen displayed exceptional intelligence and determination. She excelled academically and stood out among her peers due to her insatiable thirst for knowledge. Recognizing her potential, Carmen’s teachers encouraged her to pursue higher education despite financial constraints.

Chapter 2: Educational Journey

Despite facing various obstacles along the way, Carmen managed to secure a scholarship that allowed her to attend a prestigious university in Mexico City. She pursued a degree in Business Administration with a specialization in Finance. During her time at university, she exhibited exceptional leadership skills by actively participating in extracurricular activities and organizing events that aimed to empower women within the male-dominated field of business.

Driven by an insatiable desire for knowledge and personal growth, Carmen decided to broaden her horizons by pursuing postgraduate studies abroad. In 1990, she was accepted into Harvard Business School’s renowned MBA program on a full scholarship.

Chapter 3: Breaking Barriers in the Corporate World

Armed with a world-class education and an unwavering determination to succeed, Carmen embarked on her professional journey. She faced numerous challenges and prejudices as a woman of color in the corporate world, but she refused to be defined by societal expectations.

Carmen’s career took off when she joined a leading financial institution as an analyst. Her exceptional analytical skills and ability to navigate complex financial landscapes quickly earned her recognition within the company. Despite encountering skepticism and prejudice from some colleagues, Carmen remained steadfast in her pursuit of excellence.

Chapter 4: Entrepreneurial Ventures

Carmen’s entrepreneurial spirit was evident from an early age. In 1995, after gaining considerable experience in the corporate sector, she decided to take a bold leap and establish her own company. Carmen founded Pacheco Enterprises, a business consultancy firm that aimed to provide expert advice and support to budding entrepreneurs.

Under Carmen’s leadership, Pacheco Enterprises flourished into a highly sought-after firm renowned for its innovative strategies and commitment to client success. Carmen’s dedication to fostering diversity within her team allowed her venture to thrive in an industry dominated by male leaders.

Chapter 5: Philanthropy and Social Impact

Deeply aware of the importance of giving back, Carmen dedicated herself not only to building successful businesses but also making a positive impact on society. She became actively involved in various charitable organizations focused on providing educational opportunities for underprivileged children.

In 2007, Carmen established the Pacheco Foundation with a vision of empowering youth through education and mentorship programs. The foundation aimed to bridge the gap between disadvantaged communities and quality education by providing scholarships, mentorship opportunities, and resources necessary for academic success.

Chapter 6: Overcoming Adversity

Throughout her life journey, Carmen faced several personal hardships that tested her resilience. In 2012, she was diagnosed with breast cancer—a battle that required immense strength both mentally and physically. Carmen’s unwavering determination and the support of her loved ones allowed her to triumph over this challenging period in her life.

Chapter 7: Awards and Recognition

Carmen Pacheco’s remarkable achievements did not go unnoticed. Her exceptional leadership, philanthropic endeavors, and contributions to society garnered widespread recognition. In 2015, she was honored with the prestigious “Business Leader of the Year” award for her outstanding accomplishments in the corporate world.

In addition to professional accolades, Carmen was also recognized for her philanthropic efforts. She received numerous awards for her dedication to educational initiatives and empowering youth from disadvantaged backgrounds.

Epilogue:

Carmen Pacheco’s journey serves as an inspiration to individuals worldwide. Her unwavering determination, resilience in the face of adversity, and commitment to making a positive impact on society have left an indelible mark on the world.

Today, Carmen continues to be actively involved in various charitable endeavors while serving as a mentor and role model for aspiring entrepreneurs. She remains a symbol of empowerment and proof that with determination and hard work, anything is possible.

Unstoppable: The Extraordinary Life of Carmen Pacheco is a testament to the power of resilience, grit, and unwavering commitment in overcoming obstacles and achieving greatness.

Celebrating National Hispanic Heritage Month

HISPANIC HERITAGE MONTH—Here at ILIO, we’d like to recognize National Hispanic Heritage Month by highlighting the deep and broad Hispanic cultural influence on music. The artists chosen here are but a few who have made monumental contributions to the world’s musical treasury. Throughout every genre, there are spectacular Latin contributors. So read on, and listen to our mix with songs from such varied artists as Celia Cruz, Rosalía and many more!

Listen on:

Celia Cruz emerged during the 1950s as a Guarachas singer. She is of Cuban descent and became recognized as the queen of salsa. Her career was lauded amongst the music industry and her accolades include multiple Grammy and Latin Grammy awards. We feature “Quimbara,” which is her song that features Johnny Pacheco. Salsa brings together three distinct styles of Cuban dance, mambo, pachanga, and rumba.

The world of Latin dance music went through an evolution as it approached the modern era. In the 1980s a band called Miami Sound Machine brought new elements to this tradition. Their pop-rooted sound fused Latin dance influences with programmed drums to usher in a totally unique new sound. You can hear it on the track “Dr. Beat”.

The next realm of Hispanic music we would like to recognize is film score composition. First, Ernesto Cortázar is a Mexican composer who scored hundreds of films over the course of his career. Listen to the brilliant writing from this composer in the piece “Windmills of My Mind”. Next, Argentinian Astor Piazzolla composed for many films and ensembles throughout the world. He has a distinct sound that was defined as Nuevo Tango. This has a brilliant affect that evokes both jazz and salsa counterpoint. It can be heard on the track “Libertango”.

The jazz world was graced with the beautiful presence of Arturo Sandoval, who started recording and touring in the late 1970s. He was mentored by Dizzy Gillespie and has been embraced as one of the best jazz trumpeters in the world. You can hear his tribute to Dizzy on a rendition of “Salt and Peanuts!”. Moving into contemporary jazz, Miguel Zenon is a Puerto Rican saxophonist and composer. His style of jazz is very modern with linear improvisational stylings. “Quite de la Via, Percico” displays these skills brilliantly.

Recently, Latin acts such as Rosalía took contemporary pop music at large by storm. Rosalía has solid roots in Spanish folk music. Hailing from Spain, her eclectic sound ranges from flamenco to pop to reggaeton. She has won eight Latin Grammys and one Grammy award. Everyone from Selena to Shakira paved the way for her sound and audience.

Hispanic heritage has played a formative role in the world of music. Spanning throughout all genres, its influence and reach are unmatched. It is a privilege for us to recognize their contributions to music through this playlist. You can find it now on Spotify and Apple Music.

ComicList: New Comic Book Releases List for Wednesday, September 28, 2022, © 1995-2022 Charles S. LePage. This work is licensed under a Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International License. Any questions regarding the use of this work should be directed to Charles LePage at [email protected].

PLEASE NOTE- Not all of these titles will arrive in all stores. Let me know if any of the names or numbers are wrong. Items with release dates not verified by Diamond Distribution are individually noted.

The mean library insert size for the WGBS was 195.9 bp (SD 8.3); mean coverage was 20.2 (2.1); and the mean GC content was 22.7 (0.2). We compared methylation differences from the late exposed (LE) versus control dataset to the methylation differences in the early exposed (EE) versus control dataset to determine the role of the 56-day wash-out period on sperm DNA methylation changes. No CpG sites remained significant following conservative Bonferroni correction, so we imposed a methylation difference threshold on the top 10 K sites, retaining only those with a > 10% methylation difference in the LE relative to control datasets. This resulted in 3321 nominally significantly differentially methylated CpG (dmCpG) sites. We then analyzed methylation at those same 3321 dmCpG sites in the EE relative to control dataset. Regardless of the exposure timing, the direction of methylation change at these sites is largely the same. Linear regression of the data showed significant correlation between LE and EE mean methylation differences relative to controls (Fig. 1A, p  < 0.0001, R² = 0.82).

We separated the data based on the direction of methylation change present at the 3321 CpG sites from the LE compared to control dataset. This resulted in 1844 hypomethylated CpG sites and 1477 hypermethylated CpG sites in the exposed animals relative to the controls. Comparison of the methylation change at each CpG site shows that the magnitude of the methylation difference relative to controls is greater in the LE sperm than it is at the same CpG sites for the EE sperm for both the hypomethylated (Fig. 1B) and hypermethylated CpGs (Fig. 1C).

The mean methylation difference relative to controls across the 1844 hypomethylated CpG sites was 15.5% in the LE dataset which contrasts significantly with the 8.5% difference at the same CpG sites in the EE dataset (Fig. 1B, p < 0.0001). Similarly, there was a 13.9% mean methylation difference relative to controls across the 1477 hypermethylated CpG sites in the LE dataset as compared to a 7.5% difference at the same CpG sites in the EE dataset (Fig. 1C, p < 0.0001). These differences remained significant following Bonferroni correction for the number of CpG sites analyzed.

Ingenuity Pathway Analysis (IPA) software was used to interrogate the genes associated with the top dmCpGs from both datasets. Briefly, for those CpG sites with a greater than 10% methylation difference relative to controls, an unadjusted p-value threshold of 5.0X10⁻⁵ was implemented in both the LE and EE datasets to restrict the number to those with the greatest significance. This resulted in a list of 744 CpG sites from the LE dataset and a distinct list of 317 CpG sites from the EE dataset. Notably, only five CpG sites were in common between these two lists. IPA recognized and included 492 and 203 LE and EE dataset genes, respectively, in the downstream analyses.

We began with the IPA-generated list of 500 significant functions categories and their associated annotations for the LE dataset. Given the overrepresentation of background information associated with cancer in the IPA Knowledge Base, we removed categories associated with cancer from our analysis, as has been reported previously [19]. We also removed terms associated with only a single gene. This left 289 categories and their associated disease or function annotations. We then analyzed the data by first looking at categories with ten or more annotations (Fig. 2A, top). These categories included “cardiovascular system development, function, and disease”, “embryonic development”, “nervous system development, function, and disease”, and “developmental disorder”. We also identified the top ten most significant disease or function annotations (Fig. 2A, bottom).

From both the LE and EE datasets, distinct genes of interest were identified for independent validation. Sites were selected based on > 10% methylation difference, having ≥ 2 significantly dmCpG sites within a 100 bp region, the associated gene having a role in early life developmental processes, and the CpG site being located within the body of the gene. The following genes were selected: Homeobox Protein Hox-B9 (Hoxb9), Methyltransferase Like 11B (Mettl11b), Slit Guidance Ligand 2 (Slit2), LDL Receptor Related Protein 1 (Lrp1), Citron Rho-Interacting Serine/Threonine Kinase (Cit), Synaptotagmin (Syt17), Synaptonemal Complex Protein 3 (Sycp3), Gamma-Aminobutyric Acid Type A Receptor Subunit Beta2 (Gabrb2), Oncostatin M (Osm), 2-Phosphoxylose Phosphatase 1 (Pxylp1), Glutamate Ionotropic Receptor NMDA Type Subunit 2A (Grin2a), Synapsin III (Syn3), Netrin G1 (Ntng1), and Metastasis Suppressor 1-Like Protein 1 (Mtss1l). We confirmed performance of each of the pyrosequencing assays using defined mixtures of unmethylated and methylated DNAs (R² = 0.85–0.99, p < 0.0001–0.025, Additional file 1: Fig. S1).

Two-tailed t-tests for Osm, Gabrb2, Pxylp1, Ntng1, and Mtss1l, confirmed significant methylation changes relative to controls by pyrosequencing, in the same direction as the WGBS for the LE (Fig. 3A-E top, p < 0.05–0.005). We then broadened our analysis to assess methylation at all the CpG sites captured by the pyrosequencing assay to see if CE exposure similarly affected neighboring sites. We included sperm from both exposure groups to determine how the timing of the exposure impacted DNA methylation changes. A two-factor ANOVA—one factor being exposure status and one being CpG site—showed a significant main effect of exposure for Osm, Gabrb2, Pxylp1, Ntng1, and Mtss1l (p < 0.0001 for all genes, Fig. 3A-E bottom). Post hoc tests revealed no additional significant methylation changes for Osm.

To determine whether methylation changes in sperm of CE-exposed males were detectable in the next generation, we analyzed F1 tissues for methylation at CpG sites that were validated by pyrosequencing. We first examined Pxylp1 in F1 sperm by pyrosequencing of both exposure groups given the abundant levels of this gene’s protein product in the epididymis and in mature spermatids [25]. A two-factor ANOVA revealed a significant effect of exposure on DNA methylation in F1 sperm (p < 0.0001). Post hoc analysis revealed a significant loss of methylation in F1 sperm from animals born to LE cannabis fathers compared to controls (p < 0.05, Fig. 4A), and the difference approached significance when comparing the F1 sperm from EE and control offspring (p = 0.06).

We then analyzed methylation changes at Mtss1l in offspring hippocampus (Fig. 4B) and NAc (Fig. 4C) tissues. We observed a significant effect of paternal exposure on DNA methylation in both F1 tissues (p < 0.0001). Like the methylation changes present at these CpG sites in F0 sperm, post hoc tests revealed significant losses of methylation present in EE offspring relative to control offspring. In the hippocampus, four of the nine CpG sites analyzed were significantly hypomethylated relative to controls in the EE group (p < 0.05–0.01). This included one of the CpG sites (CpG site 1) that was initially identified via WGBS as being hypomethylated in the EE paternal sperm. In the NAc, EE offspring were hypomethylated relative to controls for four of the nine CpG sites analyzed (p < 0.05). This included the other CpG site that was initially identified via WGBS (CpG site 5) as being hypomethylated in the EE paternal sperm.

Our finding that there are heritable changes in DNA methylation for Mtss1l in brain tissues prompted us to investigate whether those methylation changes are functionally related to changes in gene expression. We first examined this relationship in hippocampal tissue of the EE and control offspring at Mtss1l CpG site 1—one of the two sites initially identified via WGBS as being affected in paternal sperm. Pearson correlations showed no significant relationship between DNA methylation and gene expression for control or exposed offspring (Additional file 3: Fig. S3A). We stratified this analysis by sex (Additional file 3: Fig. S3B and S3C) given the known sex differences in hippocampal function [26, 27] and still observed no significant relationships between DNA methylation and gene expression.

In the NAc, we examined the relationship between DNA methylation at CpG site 5 (the second site initially identified via WGBS in sperm) and gene expression for the control and EE offspring. There were no significant relationships between methylation and expression when both sexes were analyzed together (Fig. 5A). However, known sex differences in the NAc [28, 29] led us to again stratify the analysis by sex. In males (Fig. 5B), control offspring showed a significant inverse methylation-expression relationship (p < 0.05, R² = 0.61), while exposed offspring had a significant positive methylation-expression relationship (p < 0.05, R² = 0.53). In females (Fig. 5C), control offspring showed a significant positive methylation-expression relationship (p < 0.05, R² = 0.52), while exposed offspring showed an inverse though non-significant relationship.

There is growing evidence associating prenatal cannabis exposure and teratologies in babies and children and early life exposure to cannabis has been associated with cardiovascular defects in epidemiologic and animal studies [14, 30, 31]. We measured heart weights and body weights of each of the F1 offspring from the EE, LE and control fathers and normalized heart weight to body weight. One-factor ANOVA revealed a significant effect of paternal exposure on offspring heart weight (p = 0.0039). Post hoc tests showed significant increased heart weight relative to controls for both the EE offspring (p = 0.0013), and the LE offspring (p = 0.0099) (Fig. 6A). Based on physiological differences in cardiovascular function and disease in males and females [32, 33], we separated this analysis by sex. A one-factor ANOVA in females (Fig. 6B) showed a significant effect of paternal exposure on offspring heart weight (p < 0.05). Post hoc tests showed significant increases in heart weights for both EE (p < 0.05) and LE (p < 0.05) offspring. In males (Fig. 6C), a one-factor ANOVA just approached significance for the effect of paternal exposure status on offspring heart weight (p = 0.05). Post hoc t-tests showed a significant increase in heart weight relative to controls only in the EE offspring (p < 0.05).

The broad goals of the present study were to characterize the effects of paternal CE exposure on the entirety of the sperm DNA methylome in a rat model of exposure, and to determine if such effects are heritable. We did not detect significant methylation differences via WGBS after implementing a conservative Bonferroni p-value threshold. This may have been due to insufficient sample size and/or insufficient coverage. As a result, we used unadjusted p values, prioritizing based on stringent thresholds to enrich for bona fide differences.

The results showed that, regardless of the timing of the exposure, most of the methylation changes occurred in the same direction in both groups relative to the controls. We next addressed the impact of the EE-specific “washout” period on the magnitude of the methylation changes present between exposed and control animals. We found that these methylation differences were dramatically diminished in the EE animals relative to the LE animals. This supports the hypothesis that exposure cessation for the duration of at least one spermatogenic cycle is effective at minimizing many of the CE-induced epigenetic effects in sperm.

However, a distinct group of CpG sites were significantly differentially methylated relative to controls in the EE dataset that were not present in the LE dataset. These results suggest that some early exposure changes persist during the wash-out period when most other changes were minimized, and that these particular changes were specific to the early exposure period. Thus, it is possible that the spermatogonial stem cells themselves were the target of this exposure and sperm derived from these particular progenitors harbor the epigenetic modifications. Any changes that may have affected only the post-spermatogonial sperm from the early exposure (and thus might resemble changes in the late exposed sperm) might no longer have been present at the time of WGBS given the resorption of sperm over the 56-day wash-out period. Importantly, if this is true, then the persistent EE-associated changes are possibly permanent, as the progenitor pool undergoes self-renewal throughout the reproductive life of a male [34]. Different epigenetic architectures and different patterns of gene expression in these cell types and phases of spermatogenesis may have further resulted in distinct CpG sites being differentially methylated.

We used IPA to determine functional relevance of the genes with the dmCpG sites. We analyzed disease and function annotations associated with the input gene lists for both datasets. There were some notable commonalities across the LE and EE analyses, including “nervous system development, function, and disease”, “developmental disorder”, and “embryonic development”. There were also notable differences. In the LE results, “cardiovascular system development, function and disease” was among the top annotations, while for the EE results there were multiple annotations associated with behavior, including spatial learning, exploratory behavior, and cognition, as well as functions associated with embryonic development including development of sensory organ, formation of eye, and lens formation.

It is interesting that, regardless of the timing of the CE exposure, there were shared functions of the genes that were significantly differentially methylated. That we saw significant terms associated with early developmental processes supports our previous findings from studies assessing the effect of THC exposure on sperm DNA methylation [18]. In that study, we found significant KEGG pathway terms associated with early developmental processes including the hippo signaling pathway. Further, we previously provided initial evidence in support of the hypothesis that genes involved in early developmental processes are increasingly sensitive to environmentally induced epigenetic disruptions [35]. The IPA findings from this study provide additional evidence in support of this hypothesis.

We next sought to validate methylation changes at select genes identified via WGBS using bisulfite pyrosequencing. At those genes that did validate WGBS findings, we asked whether methylation changes were heritable. Whether DNA methylation changes in sperm are heritable and detectable in the next generation is one of the most critical and yet incredibly challenging questions to address. We assume that a subset of sperm is randomly impacted by preconception cannabis exposure, since methylation changes in the sperm population are not universal at any given CpG site. For detection in the next generation, one of the affected sperm must successfully fertilize the egg. Further, the methylation change(s) would need to resist post-fertilization reprogramming to persist in the developing embryo where the change is then expected to be propagated across the three germ layers. This also requires that the exposure-related methylation change does not impede the sperm’s ability to successfully fertilize an egg or the viability of the embryo in order to be detectable. Of particular relevance, studies from De Domenico [36] and Innocenzi [37] have demonstrated that in vivo exposure to the synthetic cannabinoid receptor agonist JWH-133, induced altered DNA methylation at paternally imprinted genes in sperm. These changes were subsequently detectable in placental tissues and were associated with placental and embryonic defects [36, 37].

We examined F1 sperm for Pxylp1 methylation changes given the high levels of expression of this gene in the epididymis and its enhanced presence in mature spermatids [25]. Pxylp1 is involved in the synthesis of mature glycosaminoglycan chains and possesses hydrolase and phosphatase activity [38]. It has been associated with rare diseases such as nemaline myopathy, a hereditary congenital neuromuscular disorder characterized by muscle weakness, impaired speech abilities, and hypoventilation [39]. There was significant hypomethylation in sperm from the F1 offspring of the LE animals compared to controls; a methylation change that remarkably resembles that observed in the F0 LE paternal sperm. These data are consistent with intergenerational transmission of this paternal preconception exposure. It further demonstrates that the F1 generation’s sperm population is primed to pass this epigenetic alteration to the F2 generation, indicating the potential for transgenerational inheritance.

Given the involvement of Ntng1, Gabrb2, and Mtss1l in neuronal processes, we examined F1 hippocampal and NAc tissues to determine if the sperm methylation alterations are transmissible to the F1 generation. We observed changes in Mtss1l DNA methylation in both the F1 hippocampus and NAc comparable to the father’s sperm, consistent with intergenerational transmission of this epigenetic effect. There was an interesting temporal observation for this gene, where in both generations, only the EE fathers and their offspring exhibited methylation changes while there were no meaningful changes in the LE paternal sperm or F1 tissues. The most parsimonious explanation for these results is that the F0 spermatogonial stem cells were altered, since these changes were only evident in the exposed fathers in which there was sufficient time for the sperm derived from the spermatogonia to have been produced and matured. Methylation changes present in their offspring, led us to next assess the relationship between DNA methylation and Mtss1l gene expression in the F1 brains.

Analysis in the hippocampus by F0 exposure group did not reveal significant relationships between methylation and expression, and no meaningful differences were found between offspring of exposed versus control animals. However, there are sex differences in hippocampal function, which prompted us to separate male and female offspring for analysis. Control males showed a positive correlation between methylation and expression, while in the females, there was an inverse relationship. Interestingly, paternal preconception exposure to cannabis—followed by a wash-out period—caused these relationships to reverse, where male offspring now displayed an inverse relationship and females had a positive relationship between methylation and expression. These relationships were not statistically significant; however, given that Mtss1l plays a role in regulating synaptic plasticity, even slight methylation-mediated changes in expression could have important functional consequences. Indeed, we have previously shown that a 1% change in methylation can double or halve the level of transcription of the gene encoding IGF2 [40].

The NAc and hippocampus exhibited similar patterns of methylation-expression relationships. When we stratified analysis by sex, control males had an inverse methylation-expression relationship that was significant, while control females had a significant positive relationship. Paternal preconception cannabis exposure resulted in a significant positive Mtss1l methylation-expression relationship in the male EE offspring in the NAc, and an inverse relationship in the EE female offspring. This was evident despite the wash-out period following the paternal exposure.

Literature on Mtss1l is sparse, but recent work has focused on the role of this I-BAR domain-containing gene in regulation of plasma membrane dynamics, dendritic spine formation, and postsynaptic currents [41]. Studies have demonstrated that exercise alters expression regulation of Mtss1l and have identified this gene as a novel effector involved in synaptic rearrangement in an exercise-dependent manner [41]. The ability of the Mtss1l function to be influenced by exercise raises questions about its vulnerability to other types of influences, as we have found here with cannabis. In other studies, Mtss1l mutations have been associated with severe neurodegeneration and brain iron accumulation (NBIA) [42]. While this area of research is still growing, the role of epigenetics in this syndrome is not yet understood [42]. However, abundant literature suggests a role of the environment in neurodegenerative disorders, which may be mediated in part via epigenetic mechanisms [43]. There is also a role for mitochondrial dysfunction in NBIA [42]. This presents another possible avenue for future study with regard to epigenetics, since cofactors necessary for the proper establishment and maintenance of the epigenome are supplied by the mitochondria [44].

We were especially intrigued by the effect of paternal preconception CE exposure on sex-specific relationships between Mtss1l DNA methylation and gene expression. Increasing numbers of studies report sex differences across the epigenome, particularly in brain tissues [45,46,47,48,49]. Epigenetic mechanisms play important roles in maintaining these innate sexual dimorphisms, and disruption of these epigenetic regulatory states can alter brain development and sexualization [50]. Rat models of early life exposure to bisphenol A have shown altered brain sexualization [50], and preconception and perinatal exposures to multiple chemicals and drugs have sex-specific behavioral effects on offspring [51]. In fact, paternal exposure to CE results in impaired working memory in female but not male offspring, and decreased attention behaviors in males but not females [24]. Though we did not previously know if there are inherent epigenetically regulated sexual dimorphisms in these regions of the brain that regulate Mtss1l expression, it was a surprising finding to see the relationships between methylation and expression become inverted and reflective of the normal patterns seen in the opposite sex, as a result of paternal preconception cannabis exposure.

The final endpoint that we assessed in this study was physical abnormality in F1 offspring. Studies have demonstrated an association between perinatal cannabis use and cardiovascular teratologies in offspring [14, 30, 31]. One Hawaiian study found prenatal cannabis exposure was associated with multiple cardiovascular defects in babies [30]. An epidemiologic study by Wilson et al. found that paternal cannabis use is associated with a congenital heart defect in which the positions of the aorta and pulmonary artery are interchanged in offspring [31].

We observed a significant increase in heart weights after adjustment for body weight of offspring born to fathers exposed to CE prior to mating. This effect was significant for offspring born to both EE and LE fathers for females. Surprisingly in males this phenotype was only significant in offspring born to the EE fathers. While we do not understand the mechanism, this cardiomegaly finding is in line with studies associating paternal preconception and perinatal cannabis exposures with cardiovascular defects in offspring. Future studies examining morphological, histological, and expression changes in heart tissues are needed to confirm these findings.

It is curious that our cardiovascular findings and the heritable changes in DNA methylation and gene expression were mostly in offspring of fathers exposed to CE with a 56-day wash-out period prior to mating. This raises important questions about the mechanisms underlying heritability—whether attributable to DNA methylation alone, or if other epigenetic mechanisms might also be at play. One study that might provide additional clues examined paternal preconception exposure to stress, with one week or 12 weeks between the end of the stress and the initiation of mating [52]. Authors found that offspring neurodevelopment and adult stress reactivity were impacted by paternal preconception exposure to stress when there was a 12-week recovery period following stress, compared to those animals born to mice with just a one-week recovery [52]. The authors further demonstrated that this was mediated by stress-induced changes present in the epididymal epithelial cell extracellular vesicle cargo, including noncoding RNAs, that influence sperm maturation [52]. Thus, future studies of paternal preconception exposure to cannabis should investigate the interactions between DNA methylation and other epigenetic mechanisms, as well as extracellular vesicles, and how this might influence offspring outcomes.

One additional mechanism that should be investigated in future studies is the role of the placenta in mediating the effects of paternal exposure on offspring epigenetic and phenotypic outcomes. Gene expression in the placenta is enriched for paternally expressed imprinted genes, whose methylation may be altered following a paternal exposure such as CE [53]. Given that the placenta plays critical roles in regulating fetal growth and development and that the placental methylome changes in response to environmental cues [54], the interaction between the sperm epigenome and the placental epigenome should be investigated and any downstream effects on fetal developmental processes should be studied further.

Our study has several limitations. First, sample sizes were somewhat small for our paternal exposure model—there were 12 animals per group. Second, it is possible that a 56-day wash-out period was not sufficient for the effects of the CE exposure to be fully resolved. Longer wash-out periods should be examined to determine epigenetic mitigation effects. Third, while we targeted 60X coverage for our WGBS, the effective coverage averaged 20X for our study. Thus, we may not have had sufficient depth to detect all meaningful changes or to achieve significance following Bonferroni correction. Future work is needed to investigate mechanisms underlying the observed cardiomegaly phenotype, and to determine whether any of the epigenetic and gene expression changes observed in offspring correspond to any changes in offspring neurodevelopment or behavior.

Diego Pacheco has signed a multi-fight promotional deal with Eddie Hearn and Matchroom – and the Super-Middleweight sensation headlines Matchroom’s latest fight night in Mexico, defending his WBO International and USWBC titles against Manuel Gallegos on Friday July 7 in Monterrey, live worldwide on DAZN – with co-main action provided by Matchroom’s latest signing, Super-Featherweight star Rocky Hernandez, who takes on unbeaten Venezuelan Joniker Tovar.

Pacheco (18-0 KOs 15 KOs) turned pro with Hearn in December 2018 and the rangy Super-Middleweight has grown into one of the hottest properties in the sport, closing in on the big-guns at 168lbs and doing so with his trademark fearsome power.

The 22 year old stepped through the ropes for the first time in the UK in his last outing, stopping Jack Cullen in four rounds in Liverpool – a 15th win inside the distance in his 18 fights in the paid ranks to date.

The LA talent steps through the ropes for the seventh time as a pro, and does so over ten rounds against Gallegos (19-1-1 16 KOs) the #2 ranked 168lber in Mexico behind Canelo Alvarez, who is ranked #11 in the WBO. The Mexican will be the latest to try to halt Pacheco’s rise, and the LA native is thrilled to be continuing his journey with the global leader in boxing and expects that ride to take him to World title action soon.

“I am very happy and excited to be extending me deal with Matchroom,” said Pacheco. “We have bene on the path to the top for nearly five years and I couldn’t be in a better place to achieve my goals – to become World champion, to be unified and undisputed, and to be a role model for young kids in LA and Mexico to chase their dreams.

“I’m so happy to be headlining in Mexico on July 7. It will be a good fight against the #2 ranked Super-Middleweight in Mexico – and when I win, I’ll be one step closer to getting to that World title shot.”

“I hope Pacheco is prepared to fight for his life on July 7, because I’m coming to take everything he’s got,” said Gallegos.

Pacheco’s deal comes hot on the heels of the signature of Super-Featherweight star Hernandez, and the WBC #1 ranked KO artist faces Tovar over ten rounds in Monterrey.

Hernandez (33-1 30 KOs) inked a long-term deal with Eddie Hearn today and will look to cement his status as the leading contender for the World champions at 130lbs – but unbeaten Venezuelan Tovar (21-0-1 16 KOs) says Hernandez’s bow with Matchroom will end in tears, reminding the Mexican that fellow countryman Roger Gutierrez stunned him in a round in July 2019 – the sole blemish on Hernandez’s record.

“I’m fighting another undefeated fighter to show that I’m the number one Super-Featherweight in the world and to show my promoter they made the right choice,” said Hernandez. “I’m here for 10 rounds but if he goes before then he goes!”

“”We all saw what happened the last time Rocky fought a Venezuelan,” said Tovar. “This fight will be the exact same result. I have been waiting for an opportunity to showcase my talents on a big stage, and now it presents itself. I want to thank my promoter Mr. Shane Shapiro of Shapiro Sports & Ent., Mr. Eddie Hearn of Matchroom Boxing, and DAZN for giving me this massive moment.”

“What a great day for Matchroom and DAZN in America and Mexico – securing two of the best young fighters out there and putting them together on a card in Monterrey,” said Hearn.

“Diego is the best young talent in the game – and we are delighted to have extended his deal with us. From the day he first put pen to paper with us in October 2018, there’s been no doubt in my mind that he would get to the very top.

“We’ve watched Diego blossom in and out of the ring; a powerful puncher with venom and a humble young man with his community and family in his heart – Diego can become a massive star in America and Mexico, and we are with him every step of the way.

“Rocky can almost reach out and touch that World title shot. This one is slated for ten rounds, but when you look at their respective records, both guys will be looking for an early night. Joniker is dangerous and confident, but with the champions at 130lb tuned in, Rocky will want to shine, and I expect him to do just that.”

The venue in Monterrey and ticket details will be released next week.

Sales Tax – E-commerce And Small Business

If an on the internet organization has a physical place in a particular state, such as a shop, office, or storehouse, it should accumulate sales tax obligation from consumers because state. It is not needed to accumulate sales tax obligation for sales right into that state if a company does not have a physical visibility in a state. This regulation stemmed from a 1992 Supreme Court choice which proclaimed that mail-order vendors did not require to gather sales tax obligations available right into states where they did not have a physical visibility.

When an Internet seller does not accumulate it, on-line consumers that live in a state that gathers sales tax obligation are formally called for to pay the tax obligation to the state also. When buyers are called for to pay tax obligation straight to the state, it is described as “usage” tax obligation as opposed to sales tax obligation.

The income debt collector in the majority of states currently have some sort of official setting on shopping sales. Many states deal with the sales similarly they deal with mail-order sales; nonetheless, you must call your state tax obligation firm as well as request for assistance as well as info when it come to your specific service to make sure.

State federal governments as well as brick-and-mortar merchants remain in the quest for regulations to rescind the 1992 Supreme Court judgment. Losing out on tax obligation profits from on-line acquisitions is irritating for state federal governments, so anticipate adjustments in the future.

Practical instance: Suzie discovered the best birthday celebration existing for her spouse yet can not locate them in California, so she buys the present online from a present store with head office in Texas. The provider has every one of its centers in Texas as well as accumulates settlement in Texas. Suzie does not need to pay California sales tax obligation (or Texas sales tax obligation) on the present.

A couple of months later on, the online organization opens up a storage facility in California to manage its on-line orders for the whole nation. Suzie remains to purchase presents from the exact same firm in Texas however she have to currently pay California sales tax obligation.

Ought to Internet business continue to be excluded from sales tax obligations to motivate the development of on the internet service or need to it undergo the exact same taxes dealt with by store firms is not our area to choose, however it is an issue for all acquiring as well as marketing online.

If an on the internet service has a physical place in a specific state, such as a shop, service workplace, or stockroom, it should accumulate sales tax obligation from consumers in that state. If an organization does not have a physical existence in a state, it is not needed to accumulate sales tax obligation for sales right into that state. Suzie does not have to pay California sales tax obligation (or Texas sales tax obligation) on the present.

Here’s your 7-step survival guide in case you find yourself to be the last human on Earth.

So you wake up one day and find that you’re the only person left in the world. Kind of cool, right? No obligations, no rules, no limits. Go nuts! Actually, go for the exotic fruits first. If your mangos and bananas are imported from overseas, chances are they’ll be hard to get to. So eat them while you can, and enjoy every last bite. You might think that with no one stopping you, you could go anywhere! But not so fast!

Note: this is a bonus video outside of our regular uploading schedule, a recut of a previous episode we had lying around. The older version is over here in case you’re curious: https://www.youtube.com/watch?v=fdCDQIyXGnw

Watch more what-if scenarios:
Planet Earth: https://www.youtube.com/watch?v=_-HhCwYD7rc&list=PLZdXRHYAVxTJCzxwmCq0NNpYq9N9wyb2l
The Cosmos: https://www.youtube.com/watch?v=gfuJyVkMH_g&list=PLZdXRHYAVxTJno6oFF9nLGuwXNGYHmE8U
Technology: https://www.youtube.com/watch?v=CS3bBO05fpU&list=PLZdXRHYAVxTIeRY3JtgXgoGqSEB7kDdKO
Your Body: https://www.youtube.com/watch?v=QmXR46TrbA8&list=PLZdXRHYAVxTJNsV9FFeNAKl2ySsHj8GZO
Humanity: https://www.youtube.com/watch?v=fdCDQIyXGnw&list=PLZdXRHYAVxTIFnvmOeWbv-Mt8zFxSCSvZ

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“Imagination will often carry us to worlds that never were. But without it, we go nowhere.” — Carl Sagan

Produced by Underknown in Toronto, What If is a mini-documentary web series that takes you on an epic journey through hypothetical worlds and possibilities. Join us on an imaginary adventure — grounded in scientific theory — through time, space and chance, as we ask what if some of the most fundamental aspects of our existence were different.
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1 Introduction

A range of hormones, including steroid and fatty acid derivative hormone, are classified as lipid hormone because of their lipid chemical makeup. Sex hormone, adrenocortical hormone and vitamin D (VD) are three primary subtypes of steroid hormones. The three primary types of fatty acid derivative hormones are prostaglandin, leukotriene, and iso-prostaglandin. Although the human body contains relatively little of these compounds, they can have a big physiological impact such as regulating menstruation (Critchley et al., 2020), preventing anxiety (McHenry et al., 2014) and regulating blood sugar levels (Vargas et al., 2020). Hormonal imbalances can be harmful to human health (Adegoke et al., 2021). Levels of all lipid hormones are significant diagnostic indicators for possible disease status (Pelizzaro et al., 2021). Thus, early detection by measuring the level of hormone has important significance for the identification of human diseases and the advancement of medical science.

Currently, the traditional analysis methods of lipid hormone are high-performance liquid chromatography (Ozgocer et al., 2017), liquid chromatography-mass spectrometry (Zhang et al., 2021), chemical luminescence method (Abdulsattar and Greenway, 2019) and enzyme-linked immunosorbent assay (Yin et al., 2019). However, these traditional detection techniques have the limitations of complex preparation steps, high analysis costs and time-consuming. Under these circumstances, electrochemical biosensors offer certain benefits over conventional analytical techniques, including simplicity, portability and quick detection. (Felix and Angnes, 2018) which has been applied in the fields of drug research, clinical diagnosis, food safety testing, environmental monitoring and other fields (Aydin et al., 2019). The types of the electrochemical sensor of lipid hormone that have been reported including electrochemical immunosensors (Mathew et al., 2020), electrochemical aptamer sensing (Tang et al., 2022) and electrochemical molecular imprinted polymer (MIP) sensors (Rebelo et al., 2020). Many nanomaterials including graphene, carbon nanotubes, gold nanoparticles (AuNPs) and adapters were applied in the preparation of the sensor.

This paper mainly reviews the application of electrochemical biosensors in lipid hormone detection in the past 3 years, including various kinds of detection principles, the application of nanomaterials, performance and advantages of these electrochemical sensors. We also summarize the future direction of electrochemical biosensors in lipid hormone detection, containing the challenges of current techniques and emerging applications.

2 Principle of electrochemical sensors for detection of lipid hormone

The biosensor is an instrument that uses immobilized bio-sensitive materials (antibodies, enzymes, receptors, cells, microorganisms, nucleic acids, etc.) as biometric elements to recognize the required target molecule and converts the analyte concentration into electrical signals for detection (Ronkainen et al., 2010). In general, an electrochemical biosensor has three components including biometric elements, sensors and electronic systems.

Nanomaterials are often applied to modify the electrode to amplify the detection signal. Commonly used nanomaterials include carbon nanomaterials (such as carbon nanotubes, carbon quantum dots and graphene), precious metal nanomaterials [such as gold (Au) and silver (Ag)], metal oxides (such as copper oxide and titanium oxide), polymer nanomaterials (such as MIP and conducting polymers) and biological nanomaterials (such as adapters) (Raza et al., 2021). Furthermore, the synergistic effect of multi-component nanomaterials can provide more obvious additional advantages.

The common detection methods of electrochemical sensors are cyclic voltammetry (CV), differential pulse voltammetry (DPV), square wave voltammetry (SWV) and electrochemical impedance spectrum (EIS). These detection methods all have their own uses and characteristics: CV may be used to verify both the reversibility of a reaction and the existence of intermediates in redox processes. With DPV and SWV, target analytes can even be detected at picomolar or femtomolar concentrations (Goud et al., 2021). EIS may be used to determine if the electrode/electrolyte interface impedance changes when the target is bound to the surface-immobilized biorecognition element. It has broad measured range and high detection stability as its main traits (Li et al., 2019). With the progress of electrochemical sensors, the detection method has gradually developed from desktop to portable and wearable biosensors (Samson and Koh, 2020). The schematic representation of the electrochemical biosensor for detection of lipid hormone is shown in Figure 1.

FIGURE 1. Schematic representation of the electrochemical biosensor for detection of lipid hormone.

3 Research on the electrochemical sensor of lipid hormone

Electrochemical sensors for the detection of lipid hormone have had significant success recently. A summary of electrochemical sensors for the detection of lipid hormone is listed in Table 1.

TABLE 1. A summary of electrochemical sensors for the detection of lipid hormone.

3.1 Electrochemical sensors for the detection of steroid hormone

Current electrochemical sensing technology to detect steroid hormone is mainly used for early diagnosis of disease, pregnancy research, food toxicity and pollution levels (Kelch et al., 2020). The following is a review of the electrochemical sensors of progesterone, 17β-estradiol, testosterone, cortisol, and VD.

3.1.1 Electrochemical sensors for the detection of progesterone

Progesterone is most commonly used as a pregnancy biomarker (Goh et al., 2016), which also has a central role in biology and medicine (Kanninen et al., 2019; Polat et al., 2020). Monitoring progesterone concentrations has been reported to be important in the autoimmune disease in women during menstruation (Chang and Wang, 2020), the dairy industry that can supply trustworthy data on mammalian reproduction by keeping track of progesterone levels (Yu and Maeda, 2017) and the environmental health protection by monitoring the progesterone in the wastewater (Cui et al., 2021).

Naderi and Jalali modified glassy carbon electrode (GCE) for progesterone accurately trace analysis in human serum samples and medicines using multi-wall carbon nanotubes (MWCNTs), AuNPs and poly-L-serine (Naderi and Jalali, 2020). DishaKumari et al. (2021) used magnetic graphene oxide (MGO) nanocomposite as an electrode material for the loading of bio-receptors. The increased surface area with strong electric conductivity improved sensor responsiveness. This design amplified the electrochemical signal and avoided the requirement of labeling enzymes and substrates.

Recently, progesterone detection has been accomplished using a variety of aptamer-based sensor designs. A photo-electrochemical sensor was created by Li et al. (2020) using magnetic-optical Fe₃O₄@SiO₂@TiO₂-NH₂-aptamer-nanoparticles. The sensor could selectively capture progesterone in complex biological samples. Samie and Arvand prepared a label-free electrochemical progesterone aptamer sensor using graphene quantum dots-NiO-Au hybrid nanofibers/NH₂ functionalized MWCNTs (GQDs-NiO-AuNFs/f-MWCNTs) and was successfully used to determine progesterone in human serum samples and pharmaceutical preparations (Samie and Arvand, 2020). Moreover, Zhu’s team simultaneously used antibodies and aptamer together to propose a progesterone sandwich assay, designing a sandwich-structured cathode photochemical biosensor (Zhu et al., 2020). The advantages of the sensor were easier progesterone determination, higher sensitivity and selectivity.

3.1.2 Electrochemical sensors for the detection of estradiol

So far, multiple electrochemical sensors of estradiol were aiming for 17β-estradiol, which is a natural estrogen secreted by humans and domestic animals with the strongest estrogen activity. Even trace amounts of exogenous 17β-estradiol entering the body can cause significant damage to our health (Pu et al., 2019). So, effective 17β-estradiol monitoring is important.

In the research field of 17β-estradiol electrochemical sensor, composite nanomaterials made of carbon nanomaterials and other nanomaterials have been widely used. Tanrikut’s team prepared a highly efficient sensor to detect 17β-estradiol by using the NiFe₂O₄-mesoporous carbon (NiFe₂O₄-MC) nanocomposite which displayed an optimal electron transfer rate (Tanrikut et al., 2020). Supchocksoonthorn et al. designed a 17β-estradiol sensor employing carbon dots/polyaniline (CDs/PANI) composite (Supchocksoonthorn et al., 2021). 17β-estradiol and CDs/PANI are connected by hydrogen bonds and stacking to govern adsorption. Chang et al. demonstrated a split aptamer-based electrochemical estradiol aptamer sensor, with the first piece functionalized with adamantane and the second piece that had gold nanoparticles label (Chang et al., 2021). The disposable laser-scribed graphene electrode strip with exceptional sensitivity was successfully created from the sensing platform. In order to find 17β-estradiol in the eel Lee et al. (2020) serum, developed a screen-printed carbon electrode (SPCE) with 17β-estradiol-imprinted poly (aniline-co-metanilic acid) (EIPs) and tungsten disulfide (WS₂). Da Silva and Pereira modified the electrode with MIP and carbon black (CB) to improve the sensitivity of the electrode by more than 173% compared with GCE (da Silva and Pereira, 2022). This electrochemical sensor could be easily fabricated and detect 17β-estradiol rapidly with a limit of detection (LOD) of 30 nM.

In addition to MIP electrochemical sensors based on carbon nanomaterials, Regasa and Nyokong created an electrochemical sensor based on MIP supported by AgNPs capped with 2-mercaptobenzoxazole (Regasa and Nyokong, 2022). The sensor was used to measure 17β-estradiol in actual water samples without the need of sample preconcentration processes, resulting in satisfactory selectivity, sensitivity, reusability and storage stability performances.

3.1.3 Electrochemical sensors for the detection of testosterone

Testosterone is the most essential steroid released by testicular stromal cells (Gugoasa and Stefan-van Staden, 2018) whose levels are related to many male hormone disorders and cardiovascular diseases.

To create a testosterone-imprinted electronically conductive polymer (TIECP) on the sensing electrodes, Liu et al. optimized the synthetic self-assembly of poly (aniline-co-metanilic acid) and testosterone using an electrochemical method (Liu et al., 2020). This technique optimized the conductivity of nanomaterials. Moreover, Bulut’s team synthesized a new phenylenediamine-benzodithiophene polymer (pBDBT) used to manufacture biosensors for testosterone detection (Bulut et al., 2020). A platform for real-time field detection was provided by glutaraldehyde-fixed testosterone antibodies on the polymer-coated SPCE surface. It can be used for testosterone analysis in illicit drugs.

3.1.4 Electrochemical sensor for the detection of cortisol

Cortisol is an important glucocorticoid found in a variety of biological fluids. Abnormally elevated cortisol levels can cause hypertension, damage to muscle tissue and immune system.

Now, more and more studies of electrochemical sensors for cortisol are being developed. For example, Vargas et al. (2020) developed a dual-electrochemical immunosensor based on gold microchip for the simultaneous detection of insulin and cortisol, which relied on competitive immunoassays with alkaline phosphatase (ALP) labeling . In addition, Liu et al. (2021) designed the electrochemical immunosensor for flexible AuNPs/MWCNTs/polydimethylsiloxane thin films and Naik et al. (2021) designed the “smart bandage” microfluidic platform sensor for graphene/silver solution. These three sensors are all used to detect cortisol and have the potential for instant applications.

In the study of cortisol molecularly imprinted polymers sensors, Tang et al. (2021). used a high permeability sweat-absorbing porous hydrogel to prepare a non-invasive, touch-based MIP electrochemical sensor Besides, poly glycidylmethacrylate-co ethylene glycol dimethacrylate (GMA-co-EGDMA) were used for the research of flexible MIP biosensor (Mugo and Alberkant, 2020). These two sensors have the advantages of being stretchable, small-portable and without sampling, and can serve as human wearable devices and instant application devices.

In addition to using a single biometric element to develop sensors, Huang et al. (2021) presented a highly sensitive and selective electrochemical sensor with an aptamer-antibody sandwich mode A specific combination of antibodies and aptamers was utilized to identify the target cortisol. Whereafter, Yu et al. (2022) suggested a novel electrochemical aptamer sensor using functionalized graphene (FG) and nitrogen-deqcarbon quantum dots (N-CQDs) integrating MIP techniques for trace analysis of cortisol in saliva samples.

3.1.5 Electrochemical sensors for the detection of VD

VD is not only a lipid-soluble vitamin but also an immunomodulatory hormone that has two present forms of VD₂ and VD₃ (Charoenngam and Holick, 2020). As the best marker for the monitoring of VD levels, 25-hydroxy VD (25(OH)D) is frequently employed in clinical diagnosis (Binkley et al., 2010; Farrell and Herrmann, 2013).

In the field of VD electrochemical sensors based on carbon nanomaterials, Thangphatthanarungruang prepared a graphene nanocomposite sensor for simultaneously measuring vitamins (A, D, E and K) that are lipid-soluble in various matrix samples (infant milk, yogurt and parsley) (Thangphatthanarungruang et al., 2020). Anusha designed a label-free impedance sensor using ethyl-3-(3-dimethyl aminopropyl) carbodiimide-N-hydroxysuccinimide/graphitic carbon nitride-β-cyclodextrin (EDC-NHS/GCN-β-CD)@AuNPs composite to assay serum samples for the presence of 25(OH)D₃ (Anusha et al., 2022). It is less destructive to its biomolecular activity, thus improving the sensitivity of detection. Kaur et al. (2021) designed a voltammetric immunosensor based on molybdenum sulfur MoS₂/AuNPs/tin fluoride oxide with 25(OH)D₃ as the target molecule The result showed that the MoS₂-modified AuNPs model demonstrated excellent detection conductivity, sensitivity and stability.

Some electrochemical sensors using VD₃ aptamer were also reported. Wadhwa’s team prepared a portable electrochemical aptamer sensor to identify VD₃ employing graphene quantum dot-gold hybrid nanoparticles (GQD-Au) with a LOD of 0.70 nM (Wadhwa et al., 2020). Subsequently, Shuo et al. (2021) proposed a novel electrochemical aptamer sensor for sensitively detecting 25(OH)D₃ by fixing DNA tetrahedra on Au surfaces and a technique for catalytic hairpin assembly (CHA) amplification.

In addition, Anusha’s group synthesized a new composite material based on polyaniline-polypyrrole (PANI-PPY) copolymer doped with silver-cobalt (Co-Ag) and ionic liquid (IL) (Anusha et al., 2021). The composite material was first used to modify the GCE. Later, a manual paper sensor was created using the suggested material. And using two different tests to detect VD₃ in serum and urine samples.

3.2 Electrochemical sensors for the detection of fatty acid derivative hormone

The fatty acid derivative hormone is a metabolite produced by lipids through oxidative and enzymatic metabolic pathways including iso-prostaglandin and prostaglandin. Prostaglandin regulates pathological processes in female reproductive function (Niringiyumukiza et al., 2018), inflammation, and tissue repair (Rael, 2016). The lipid peroxidation biomarker 8-iso-prostaglandin F2α and carotid subclinical atherosclerosis showed a significant positive connection (Alharby et al., 2019).

In previous reports, electrochemical sensors of prostaglandin E1 (Zheng et al., 2016) and 8-iso-prostaglandin F2α (Sanchez-Tirado et al., 2017) have been successively reported. With the deepening understanding of the fatty acid derivative hormone and the development of electrochemical sensors, Ganguly et al. (2021) developed a three-electrode planar gold microelectrode system with flow-based nanopore membranes for electrochemical immunosensors for detecting prostaglandin E2 (PGE2). This sensor can be used in both clinical and home settings for a more immediate, fast and accurate diagnosis of urinary tract infection.

4 Conclusion and future perspectives

This paper mainly describes the recent research progress of electrochemical biosensors in the field of lipid hormone detection in the past 3 years. This could help the future development of such sensors in medicine and science. Furthermore, combined with the important role and significance of lipid hormones such as progesterone, estradiol, testosterone, cortisol, VD, prostaglandin in regulating human life activities and disease control, and the easy application of electrochemical sensors, the development of lipid hormone electrochemical sensors has attracted more and more attention in recent years.

Overall, so far, there are more comprehensive and mature studies of steroid electrochemical sensors than those of fatty acid derivative. Current electrochemical biosensors for fatty acid derivative hormone use only antibodies as recognition elements. In the future, electrochemical sensors will have considerable potential for development and applications in the detection of lipid hormone. A key opportunity for the development of electrochemical sensor and biosensor platforms is the introduction of innovative functional nanomaterials and analytical technologies. Such as new electrode materials with more selectivity and sensitivity, more portable wearable sensors and instant application sensors still need to be continuously explored by researchers.

Author contributions

TZ: conceptualization and writing—original draft preparation. XD and ZZ: validation, writing—review and editing, and project administration. All authors contributed to the article and approved the submitted version.

This work was supported by grants from the National Natural Science Foundation of China (32170921 and 32170734), and the Young Talent of Lifting Engineering for Science and Technology in Shandong, China (SDAST2021qt07).

Conflict of interest

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Publisher’s note

All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.

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