Researchers at Jinan University in Guangzhou, China, have developed the first stem cell-based treatment strategy for androgen deficiency due to male hypogonadism, whereby adult rodent skin cells are directly converted into functional testosterone-producing Leydig-like cells.

Those newly generated cells not only survived, but also were shown to restore physiologically normal testosterone levels when they were transplanted into the testes of male rats and mice with hypogonadism, the researchers reported Dec. 22, 2016, in Stem Cell Reports.

Male hypogonadism causes reduced testosterone levels in approximately 30 percent of older men. It is characterized by symptoms including mood disturbances, sexual dysfunction, decreased muscle mass and strength, and decreased bone mineral density with increased risk of fractures.

One primary cause of male hypogonadism is dysfunction of the testicular testosterone-producing Leydig cells. While testosterone replacement therapy (TRT) can alleviate some symptoms resulting from hypogonadism, it may also increase the risk of prostate and cardiovascular problems.

Those potentially serious side effects associated with TRT prompted the researchers to develop their alternative cell-based treatment strategy for male hypogonadism.

“Long-term TRT will maintain the serum testosterone concentration at a stable level; it will be without physiological pulses, which will induce some side effects, including stimulation of prostatic growth, worsening of voiding dysfunction and even induction of prostate cancer,” co-senior study author Zhijian Su, an associate professor in the Department of Cell Biology at Jinan University, told BioWorld Today.

“Our study is the first to report a method for generating Leydig cells by means of direct cell reprogramming,” added senior study co-author Yadong Huang, a professor in the Department of Cell Biology at Jinan University.

“This alternative source of Leydig cells will be of great significance for basic research and provides the attractive prospect of clinical application in the field of regenerative medicine,” Huang said.

“We hope to be able to use these functional regenerated Leydig cells for treating people of all ages who have hypogonadism,” Su said. “We also hope this technology can play a role in female reproduction by increasing hormone levels.”

Functional Leydig cell transplantation could also be a promising alternative to hormone replacement therapy, restoring normal physiological levels of hormone for a longer period of time.

However, stem cell-based approaches are costly, time-consuming and limited by ethical concerns and the risk of tumor formation. Huang and Su therefore decided that directly converting adult skin cells into Leydig cells would be a faster, safer regenerative approach to managing hypogonadism.

To test that idea, the researchers screened 11 transcription factors that could affect the ability of Leydig cells to produce testosterone. Using lentiviral vectors to force expression of three of those transcriptional factors, known as Dmrt1, Gata4 and Nr5a1, they were then able to directly reprogram mouse skin fibroblasts into functional Leydig-like cells.

The newly regenerated cells were capable of producing robust levels of testosterone. Moreover, when transplanted into the testes of rats or mice with hypogonadism, the cells survived and were shown to be capable of restoring serum testosterone to normal physiologic levels.

“In our study, the activity of androgen synthesis in the newly generated Leydig-like cells, like natural Leydig cells, was triggered by luteinizing hormone in vitro and regulated by the hypothalamus-pituitary-gonad axis in vivo. Therefore, normal levels of serum testosterone can be maintained, including physiological pulses,” said Su.

According to the Jinan University researchers, future studies should aim to improve the efficiency of that treatment approach to generate a pure population of cells that closely mimic natural Leydig cells.

They are also examining in more detail the mechanisms underlying the direct conversion of skin cells into Leydig-like cells. In addition, they are examining direct cellular conversion strategies using small molecules and other nonviral methods to minimize the chance of adverse events, including cancer.

“Although lentiviral vectors are generally safe, it is better to avoid using such heterogenous nucleic acids, especially in the clinical setting. Small molecules that do not integrate into the host genome, but which can stimulate the steroidogenic-related genes to express the desired proteins, are preferable,” said Su.

“We are hopeful that this research will pave the way for clinical trials testing a novel regenerative medicine approach to treat androgen deficiency in men,” said Su. “While we do not currently have a detailed schedule for clinical trials using this technique, we believe they will begin in the next few years,” he said.

“Meanwhile, we are performing experiments on converting human fibroblasts into functional Leydig-like cells. We hope that we can obtain the dedifferentiated cells that have the same excellent characteristics as natural Leydig cells, in terms of their pattern of gene expression and capacity for androgen synthesis.”