Sperm cargo is richer and more important than we thought

Amazing stuff!

I remember way back then when some feminists and some female women's rights activists mocked/dismissed men as mere, dispensable sperm providers. 

"Sperm have long been thought of as streamlined DNA delivery vehicles, carrying little more than a father’s genes to the egg. But a new study shows that in mice, sperm may transmit the father’s influence in another way: During their passage through the epididymis, the coiled tube where they mature after leaving the testes, sperm pick up messenger RNAs (mRNAs)—RNA transcripts of genes that contain the genetic instructions for making proteins. And these mRNAs seem to be transferred to the fertilized egg, a team reports this week in Nucleic Acids Research.

Researchers already knew sperm ferry small RNAs, RNA fragments that can silence gene expression and have been implicated in transmitting the effects of paternal diet, stress, and  exercise to offspring. But mRNAs could be a far more direct route for paternal influence ... The work doesn’t prove these mRNAs from sperm actually function in embryos, she noted, but the discovery of unexpected cargo is “potentially very significant.”"

"... The researchers also found that some mRNAs present in mature sperm are absent from unfertilized eggs, but appear in zygotes after fertilization—suggesting sperm deliver these transcripts to the embryo, ... Because environmental conditions can trigger mRNA production in the father, the observed mRNA transfer “really establishes a mechanism for how the environment can directly influence sperm to then potentially influence the next generation,” ...

the researcher injected long RNA sequences into parthenotes—mouse eggs triggered to divide and develop without sperm. They found the injections shifted the cells’ gene expression to resemble normally fertilized embryos. (They team used RNAs longer than 200 nucleotides, but not mRNA ... The results suggest large RNAs from sperm “can do something after fertilization to regulate embryonic gene expression,” ...

The mechanism may not be limited to mice. The team also sequenced mRNAs from mature human sperm and found counterparts to many mRNAs in mouse sperm, suggesting humans may also deliver RNA messages to their offspring this way."

From the abstract:

"The epididymis plays a critical role in promoting sperm maturation, including remodeling the sperm RNA payload. While small RNAs have been extensively studied in this context, the epididymal contribution to larger sperm RNAs, such as messenger RNAs (mRNAs), remains underexplored. This is largely due to the translational quiescence of mature spermatozoa and the hypothesis that these RNAs are residual by-products of spermatogenesis.

Yet, mRNAs carried by sperm have been detected in the zygote, indicating they may act beyond fertilization. However, whether epididymal somatic cells contribute mRNAs to sperm, as they do small RNAs, has not been experimentally examined. Here, we provide a comprehensive analysis of the mRNA landscape of mouse sperm, epithelial cells, and extracellular vesicles (EVs) isolated from the proximal (caput) and distal (cauda) epididymis.

Through this analysis and sperm-EV co-incubation experiments, we demonstrate the transfer of mRNAs from epididymal EVs to sperm.

Further, through sperm RNA microinjection into zygotes, we uncover gene regulation in the early embryo driven by the introduction of sperm RNAs, specific to >200-nucleotide RNA species.

These findings reveal the dynamic mRNA profile of sperm that is delivered to the egg and demonstrate that RNA species beyond small RNAs are capable of influencing preimplantation embryo gene expression."

ScienceAdviser

Sperm carry unexpected genetic messages "Maturing mouse sperm get loaded with full-length messenger RNAs that are transferred to fertilized egg, suggesting a new route for paternal influence"

Epididymal extracellular vesicles harbor and convey mRNA to sperm for transfer to zygotes (open access) [This is a long research article!]

Graphical abstract

Study advances safe, reversible male contraceptive without hormones

Good news! Why did men have to wait for so many decades since the contraceptive pill for women came out!

"A proof of principle study in mice, six years in the making, shows how targeting a natural checkpoint in meiosis, the process by which sex cells reproduce, safely stopped sperm production. ...

The researchers made use of JQ1, a small molecule inhibitor that was developed as a research tool to study cancer and inflammatory disease. Because of its neurological side effects, it wasn't a viable therapy for disease, or as a final contraceptive, but it's known to disrupt a stage of meiosis called prophase 1. This enabled Cornell researchers to provide the first proof of principle that meiosis – and sperm production – can be targeted safely and reversibly. ..."

"... In the study, the researchers administered JQ1 in male mice for three weeks. They found that the mice produced no sperm, and that all the molecular parameters of meiosis were disrupted, including chromosomal behavior during prophase 1.

Then they stopped delivering JQ1, and within six weeks, most of the healthy parameters of prophase 1 returned, along with normal sperm production. They then bred those mice and found they were all fertile, and bred the pups to show that they too were fertile, yielding healthy offspring. ..."

From the significance and abstract:

"Significance

Few reversible male contraceptives have advanced toward clinical translation, largely because the optimal biological stage for safe intervention remains undefined.

Meiosis represents a natural checkpoint in sperm production where transient inhibition could achieve precise and reversible fertility control.

Using the small-molecule BRDT inhibitor (+)-JQ1, we demonstrate that brief suppression of meiotic prophase I halts spermatogenesis yet permits complete recovery of germ-cell differentiation, recombination fidelity, and fertility after withdrawal. While acknowledging the need for robust future safety assessments of any candidate drugs, these studies provide a blueprint for developing new contraceptive approaches that act safely and selectively within the germline.

Abstract

Developing safe, reversible, and nonhormonal male contraceptives has been hindered by the lack of defined biological windows that can be transiently interrupted without compromising long-term fertility.

Here, we tested whether meiotic prophase I can serve as such a window by pharmacologically inhibiting the testis-specific chromatin reader BRDT using the small-molecule bromodomain inhibitor (+)-JQ1 as proof-of-principle.

Short-term JQ1 administration (3 wk) selectively disrupted the pachytene transcriptional program, depleted postmeiotic germ cells, and induced a reversible arrest in spermatogenesis.

Upon drug withdrawal, prophase I cytological markers normalized within 6 wk, accompanied by restoration of testis architecture and germ-cell composition. Crossover metrics and transcriptional programs recovered more gradually, reaching full normalization by 30 wk alongside complete restoration of fertility and fecundity.

These results demonstrate that meiotic prophase I can be transiently inhibited to suppress spermatogenesis reversibly without inducing lasting genomic or reproductive defects, defining a stage-specific framework for the rational design of nonhormonal male contraceptives."

Global Health NOW

Study advances safe, reversible male contraceptive without hormones

Breakthrough takes big step toward safe, reversible male contraception

Meiotic prophase I disruption as a strategy for nonhormonal male contraception using small-molecule inhibitor JQ1 (open access)

Fig. 1 Three-week JQ1 treatment causes postmeiotic cell loss with full recovery after withdrawal.

Cross‑section of testis tubules showing stages of spermatogenesis: green meiotic cells, pink developing sperm (dots in top right corner) and pink mature sperm (long cells in middle left), and the pre-meiotic white/gray spermatogonia along the basement membrane (lower left) are stem cells the researchers aim to preserve.

Hidden evolution of genetic mutations in sperm raises disease risk for children as men age

It was known for several decades that sperm deteriorates with age, but this study provides more details and how it develops over time. Probably, older men should have their sperm checked if they intend to become fathers.

"Harmful genetic changes in sperm become substantially more common as men age because some are actively favoured during sperm production, new research has revealed. 

In a landmark study ... researchers ... have comprehensively mapped how harmful DNA changes in sperm cells can increase across the genome as men age.  ...

In a new study, researchers used NanoSeq, an ultra-accurate DNA sequencing method to analyse sperm from 81 healthy men, aged between 24 and 75 years, with unprecedented precision. The sperm samples were collected as part of the TwinsUK cohort, the UK’s largest adult twin registry, providing a unique opportunity to study genetic variation across a diverse and well-characterised population.

The results show that around 2 per cent of sperm from men in their early 30s carried disease-causing mutations, but this rises to 3–5 per cent of sperm from middle-aged (43 to 58 years) and older men (59 to 74 years). Whilst men may be more likely to father children at a younger age, the researchers found that in those aged 70, 4.5 per cent of sperm carry disease-causing mutations. ...

This risk is driven not just by the steady build-up of random changes in DNA, but by a subtle form of natural selection acting on the sperm-producing cells in the testes, giving some mutations a competitive edge during sperm production. 

The researchers identified 40 genes where certain changes in DNA are favoured during sperm production, including many linked to childhood diseases, severe neurodevelopmental disorders and inherited cancer risk. While 13 genes  have already been associated with this process, the new findings show it is far more widespread than previously understood, affecting a broad range of genes important for cell growth and development. ..."

From the abstract:

"Mutations that occur in the cell lineages of sperm or eggs can be transmitted to offspring. In humans, positive selection of driver mutations during spermatogenesis can increase the birth prevalence of certain developmental disorders. Until recently, characterizing the extent of this selection in sperm has been limited by the error rates of sequencing technologies.

Here we used the duplex sequencing method NanoSeq to sequence 81 bulk sperm samples from individuals aged 24–75 years. 

Our findings revealed a linear accumulation of 1.67 (95% confidence interval of 1.41–1.92) mutations per year per haploid genome driven by two mutational signatures associated with human ageing.

Deep targeted and exome NanoSeq5 of sperm samples identified more than 35,000 germline coding mutations.

We detected 40 genes (31 newly identified) under significant positive selection in the male germline that have activating or loss-of-function mechanisms and are involved in diverse cellular pathways.

Most of the positively selected genes are associated with developmental or cancer predisposition disorders in children, whereas four of the genes exhibited increased frequencies of protein-truncating variants in healthy populations.

We show that positive selection during spermatogenesis drives a 2–3-fold increased risk of known disease-causing mutations, which results in 3–5% of sperm from middle-aged to older individuals with a pathogenic mutation across the exome.

These findings shed light on germline selection dynamics and highlight a broader increased disease risk for children born to fathers of advanced age than previously appreciated."

Hidden evolution in sperm raises disease risk for children as men age "Researchers reveal how certain harmful DNA variations become more common in sperm as men age, raising genetic disease risk for offspring."

Sperm sequencing reveals extensive positive selection in the male germline (open access)

Fig. 1: Mutational burden and signature analysis in sperm and matched blood.

Have scientists uncovered an alternative to condoms and vasectomies?

Good news! Hopefully, finally men have better options too! However, this is very early research!

Gender equality in contraceptives!

That men have been so dependent for so many years on women to effectively control reproduction without interfering with the joy of sex was hard to swallow (pardon my pun).

"... surveys show most American men are interested in using male contraceptives, yet they have almost no options. Recent attempts to develop drugs that block sperm production, maturation, or fertilization have had limited success, providing incomplete protection or severe side effects. New approaches to male contraception are needed, but because sperm development is so complex, researchers have struggled to identify parts of the process that can be safely and effectively tinkered with.

Now, scientists at the Salk Institute in California have found a new method of interrupting sperm production that is both non-hormonal and reversible. The study, just published in Proceedings of the National Academy of Sciences under the title “Targeting nuclear receptor corepressors for reversible male contraception,” has found a  new protein complex in regulating gene expression during sperm production. The researchers demonstrate that treating male mice with an existing class of drugs called HDAC (histone deacetylase) inhibitors can interrupt the function of this protein complex and block fertility without affecting libido.  ...

Salk scientists found that for this to work, retinoic acid receptors must bind with a protein called SMRT (silencing mediator of retinoid and thyroid hormone receptors) that then recruits HDACs, and this complex of proteins goes on to synchronize the expression of genes that produce sperm. ..."

From the significance and abstract:

"Significance

With unintended pregnancies costing billions of dollars a year in the United States alone, the socioeconomic and health benefits of improved birth control cannot be overemphasized. Despite the pressing need for population control, male contraceptive options are limited. These rodent studies demonstrate reversible male contraception by targeting silencing mediator of retinoid and thyroid hormone receptors-retinoid acid receptor signaling to identify a non-hormonal approach to male contraception.

Abstract

Despite numerous female contraceptive options, nearly half of all pregnancies are unintended. Family planning choices for men are currently limited to unreliable condoms and invasive vasectomies with questionable reversibility. Here, we report the development of an oral contraceptive approach based on transcriptional disruption of cyclical gene expression patterns during spermatogenesis. Spermatogenesis involves a continuous series of self-renewal and differentiation programs of spermatogonial stem cells (SSCs) that is regulated by retinoic acid (RA)–dependent activation of receptors (RARs), which control target gene expression through association with corepressor proteins. We have found that the interaction between RAR and the corepressor silencing mediator of retinoid and thyroid hormone receptors (SMRT) is essential for spermatogenesis. In a genetically engineered mouse model that negates SMRT-RAR binding (SMRTmRID mice), the synchronized, cyclic expression of RAR-dependent genes along the seminiferous tubules is disrupted. Notably, the presence of an RA-resistant SSC population that survives RAR de-repression suggests that the infertility attributed to the loss of SMRT-mediated repression is reversible. Supporting this notion, we show that inhibiting the action of the SMRT complex with chronic, low-dose oral administration of a histone deacetylase inhibitor reversibly blocks spermatogenesis and fertility without affecting libido. This demonstration validates pharmacologic targeting of the SMRT repressor complex for non-hormonal male contraception."

Have scientists uncovered an alternative to condoms and vasectomies? - The Jerusalem Post Salk Institute scientists discover new target for reversible, non-hormonal male birth control

Salk Scientists Discover New Target For Reversible, Non-Hormonal Male Birth Control Oral administration of HDAC inhibitor blocked sperm production and fertility in mice without affecting libido

Targeting nuclear receptor corepressors for reversible male contraception (no public access)

Sperm, pictured inside the cross-sectioned tube of the epididymis, were not generated while mice took the HDAC inhibitor drug (top right), but after 60 days off the drug, spermatogenesis was recovered (bottom right). The left column shows sperm at the same time points in a mouse that did not receive the drug.

Israeli Scientists Grow Sperm On A Microchip

Amazing stuff!

"... scientists ... have developed an innovative platform to create sperm in a laboratory through a microfluidic system ...The sperm was grown on a special silicon chip developed ... A 3D system was built and integrated to allow the addition of testicular tissue cells. The chip enabled the researchers to grow cells from the testis in the microchip and add fresh cell culture media designed to support cellular growth. ..."

From the abstract:

"This research presents a novel testis-on-a-chip (ToC) platform. Testicular cells are enzymatically isolated from the seminiferous tubules of sexually immature mice, seeded in a methylcellulose gel and cultured in a microfluidic chip. ...  The cells develop into spheroids continuing to proliferate and differentiate. After seven weeks of culture the cells have over 95% viability. Confocal microscopy of the developed spheroids reveals a structure containing the various stages of spermatogenesis up to and including meiosis II: premeiotic, meiotic and post-meiotic germ cells. The spheroid structure also contains the supporting Sertoli and peritubular cells. ... As a benchmark, the ToC is compared to a conventional three-dimensional methylcellulose cell culture system in a well plate. Analysis via fluorescence-activated cell sorting shows more haploid cells in the chip as compared to the plates. ... This research opens new horizons for the study and realization of spermatogenesis in-vitro. It can also enable the implementation of microfluidic technologies in future therapeutic strategies for pre-pubertal male fertility preservation and adults with maturation arrest. Lastly, it can serve as a platform for drug and toxin testing."

Israeli Scientists Grow Sperm On A Microchip

Testis on a chip—a microfluidic three-dimensional culture system for the development of spermatogenesis in-vitro (no public access)

Functional Rat Sperm Generated from Stem Cells

It took only a decade to go from mice to rats!

"In the past decade, methods have been developed to generate germ cells from pluripotent stem cells for studies of development and in vitro gametogenesis. However, offspring from in vitro–derived germ cells has only been achieved in mice. ...
Now, rats join the club. A study published today (April 7) in Science reports that researchers have produced healthy, fertile rat offspring with sperm made from stem cells. ..."

From the abstract:

"The in vitro generation of germ cells from pluripotent stem cells (PSCs) can have a substantial effect on future reproductive medicine and animal breeding. A decade ago, in vitro gametogenesis was established in the mouse. However, induction of primordial germ cell–like cells (PGCLCs) to produce gametes has not been achieved in any other species. Here, we demonstrate the induction of functional PGCLCs from rat PSCs. We show that epiblast-like cells in floating aggregates form rat PGCLCs. The gonadal somatic cells support maturation and epigenetic reprogramming of the PGCLCs. When rat PGCLCs are transplanted into the seminiferous tubules of germline-less rats, functional spermatids—that is, those capable of siring viable offspring—are generated. ..."

Rat Sperm Generated from Stem Cells | The Scientist Magazine®

Functional primordial germ cell–like cells from pluripotent stem cells in rats (no public access)

Rat Sperm Generated from Stem Cells 10 years after the mouse

What took so long? Was this lack of interest or something more profound?

"More than a decade ago, scientists succeeded for the first time in generating primordial germ cells—the precursors of sperm and eggs—from embryonic stem cells, a process that led to functional sperm capable of producing offspring. The milestone, achieved in mice, had not been repeated in any other species since. ...
In recent years, though, [researchers] have been constructing rat mutants that have allowed them to visualize the development of primordial germ cells in vivo. Using fluorescent markers to trace the expression of genes that are key to the transition from stem cells to primordial germ cells, they have learned more about how gene expression changes over time, all of which was helpful to finally recapitulate the process in vitro ..."

"In the past decade, methods have been developed to generate germ cells from pluripotent stem cells for studies of development and in vitro gametogenesis. However, offspring from in vitro–derived germ cells has only been achieved in mice. Oikawa et al. extend this work beyond mice to a second rodent species, the rat, a leading animal model for biomedical research with many physiological similarities to humans. A stepwise protocol allows for the production of fetal stage rat germ cells that can produce viable offspring upon maturation in the testis and injection of the sperm into unfertilized oocytes. This system will allow comparative studies and enable broader execution and analysis of in vitro gametogenesis."

Rat Sperm Generated from Stem Cells | The Scientist Magazine®

Functional primordial germ cell–like cells from pluripotent stem cells in rats (no public access)