Bitter taste receptors: new players in female and male fertility

The mammalian taste perception provides valuable information about the nature and quality of food. Taste transduction involves the interaction of molecules (i.e. tastants) with taste receptor (TASRs) whose expression isn’t limited to the oral cavity; indeed, many studies showed how these receptors can be found also in extraoral location, wherein their function appears less obvious and remains to be understood in most cases. The expression of these receptors outspreads also to brain, respiratory tract, urinary bladder and testis, where some of these receptors are expressed in different stages of spermatogenesis. Moreover, the expression of selected member of different receptor families in ejaculated human sperm has been reported. These findings suggest a possible involvement of TASRs in sperm maturation as well as in sperm chemotactic activation and fertilization. To the best of our knowledge, no data are available regarding the expression of TASRs in the mammalian oocyte nor in the somatic cells of the follicle. Therefore, to investigate the possible role of TASRs in male and female reproductive system, focusing on their impact on human fertility, we analyzed the expression of some members of taste 2 receptors family (TAS2Rs) genes in human germ cells and follicular cells, namely granulosa and cumulus cells. The expression of genes involved in taste receptors signaling cascade was also investigated, both in female and male. Immunofluorescence analysis and western blot of TAS2Rs and related signaling pathways also carried out highlighting the expression of selected TAS2Rs genes in ejaculated sperm and in testis and, for the first time, in somatic cells from human ovarian follicle. An interesting finding of this thesis work is the demonstration of TAS2R14 as the most expressed gene in both granulosa and cumulus cells. These results suggest a key role of this receptor and also for its potential involvement in follicular cells physiology. The possible role of this receptor in reproduction is also supported by a study showed TAS2R14 to be correlated with sperm progressive motility. In order to elaborate an enrichment analysis to summarize functional information about the investigated proteins and to predict possible associations among them, a protein network analysis was performed by MetaCore software. Even if further studies are needed to better understand the precise functional role oftaste receptors in reproduction, this work try to improve our knowledge of the biological role of TAS2Rs for human male and female fertility

Increased expression of neurogenic factors in uterine fibroids

STUDY QUESTION: Are selective markers for the neuronal differentiation such as microtubule-associated protein 2 (MAP-2) and synaptophysin (SYP) as well as the nerve growth factor (NGF) expressed by fibroids, myometrium and eutopic endometrium? SUMMARY ANSWER: Neuronal markers NGF, MAP-2 and SYP are highly expressed in fibroids compared with matched myometrium, and this neurogenic pathway is upregulated by tumor necrosis factor (TNF) alpha in cultured smooth muscle cells (SMCs). WHAT IS KNOWN ALREADY: Uterine fibroids or leiomyomas are the most common benign tumors, accounting for approximately one-third of hysterectomies. The present trend is to improve the medical treatment avoiding surgery, also for fertility sparing; hence, the pathogenic mechanisms are investigated, aiming to develop new therapeutic strategy. STUDY DESIGN, SIZE, DURATION: This laboratory-based case-control study is focused on fibroids and myometrial specimens obtained between 2015 and 2017 from 15 women of reproductive age at the proliferative phase of the menstrual cycle. Leiomyomas, matched myometrium and endometrium from each woman were analyzed. Control endometrium was obtained from women undergoing surgery for ovarian cyst (n = 15). PARTICIPANTS/MATERIALS, SETTING, METHODS: qRT-PCR, western blotting and immunostaining were applied to evaluate the expression of neurogenic markers; the effects of TNF on NGF, MAP-2 and SYP expression in cultured SMCs from leiomyomas and matched myometrium were analyzed. MAIN RESULTS AND THE ROLE OF CHANCE: qRT-PCR analyses using tissues from clinical patients showed that the levels of NGF, MAP-2 and SYP mRNA were significantly higher in uterine leiomyomas compared with their matched myometrium (P < 0.05), whereas only NGF was significantly increased in eutopic endometrium compared with healthy endometrium. In primary SMCs, isolated from fibroids or from the adjacent myometrium, NGF, MAP-2 and SYP mRNA expression were significantly increased by TNF treatment (P < 0.05). Finally, human endometrial stromal cells prepared from the endometrium of patients affected by uterine fibroids display higher TNF expression (P < 0.001). LIMITATIONS, REASONS FOR CAUTION: qRT-PCR analysis and immunofluorescence validation are robust methods demonstrating a clear upregulation of neurogenic factors in leiomyomas, even though additional studies are needed to establish a correlation between increased neuronal gene expression and degree of pain, as well as the involvement of inflammation mediators in the development of the neurogenic unhinge. Therefore, more in vivo studies are needed to confirm the results achieved from primary cultured SMCs. WIDER IMPLICATIONS OF THE FINDINGS: The increased expression of neurogenic factors in uterine fibroids and endometrium may contribute to explain the painful stimuli. Accordingly, these neurogenic pathways may represent potential therapeutic avenues to treat the fibroid-related disorders. STUDY FUNDING/COMPETING INTEREST(S): This study was supported by research grants from the University of Siena. The authors declare no conflict of interest. TRIAL REGISTRATION NUMBER: N/A. © 2019 The Author(s) 2019. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserve

Characterization of Enzyme-Linked Immunosorbent Assay (ELISA) for Quantification of Antibodies against <i>Salmonella</i> Typhimurium and <i>Salmonella</i> Enteritidis O-Antigens in Human Sera

Nontyphoidal Salmonella (NTS) is a leading cause of morbidity and mortality caused by enteric pathogens worldwide in both children and adults, and vaccines are not yet available. The measurement of antigen-specific antibodies in the sera of vaccinated or convalescent individuals is crucial to understand the incidence of disease and the immunogenicity of vaccine candidates. A solid and standardized assay used to determine the level of specific anti-antigens IgG is therefore of paramount importance. In this work, we presented the characterization of a customized enzyme-linked immunosorbent assay (ELISA) with continuous readouts and a standardized definition of EU/mL. We assessed various performance parameters: standard curve accuracy, dilutional linearity, intermediate precision, specificity, limits of blanks, and quantification. The simplicity of the assay, its high sensitivity and specificity coupled with its low cost and the use of basic consumables and instruments without the need of high automation makes it suitable for transfer and application to different laboratories, including resource-limiting settings where the disease is endemic. This ELISA is, therefore, fit for purpose to be used for quantification of antibodies against Salmonella Typhimurium and Salmonella Enteritidis O-antigens in human samples, both for vaccine clinical trials and large sero-epidemiological studies

Follicular microenvironment: Oxidative stress and adiponectin correlated with steroids hormones in women undergoing in vitro fertilization

Research has been focused on determining the follicular microenviroment produced by the theca and granulosa cells since the molecular characterisation of this body fluid could lead to the understanding of several fertility problems. Oxidative stress may be one of the factors involved in female infertility since it plays a key role in the modulation of oocyte maturation and finally pregnancy. An increase in oxidative stress is correlated with inflammation and intense research was developed to understand the interaction between inflammation and adiponectin, based on the fact that many adipokines are inflammation related proteins linked to reactive oxygen species production. The aim of this study is to investigate the correlation between total adiponectin levels and oxidative stress amount in the serum and follicular fluid (FF) of women who undergone in vitro fertilization. Moreover we verified the expression of adiponectin in granulosa and cumulus cells. To clarify the predictive value of steroid hormones in human assisted reproduction, twelve steroid hormones in FF and serum, were quantified in a single run liquid chromatography/mass spectrometry, by using a multiple reaction monitoring mode and we related the serum and follicular fluids adiponectin levels with the concentration of the investigated steroid hormones