Reproductive physiology of the female cat

Ovarian cyclicity induces considerable changes in morphology, physiology and function of the reproductive organs in animals. This thesis aimed to study reproductive physiology of the female domestic cat, focusing on the cervix, uterus and uterine tube during different stages of the oestrous cycle, after mating and under pathological conditions. Transcervical catheterisation was performed using a specially designed catheter. Patency of the cervix and uterine motility were studied with the aids of fluoroscopy and scintigraphy by depositing contrast fluids and radiopharmaceutical medium in the cranial vagina. The relationship between cervical patency and oestrous behaviour, cornification of the vaginal cells and serum concentrations of oestradiol-17β were evaluated. Hysterography, a positive contrast study, was performed to illustrate the uterine appearance during the oestrous cycle in relation to histological characteristics of the endometrium. Immunohistochemistry using proliferating nuclear antigen (PCNA) was introduced to identify mitotic activity of the endometrial cells. Patency of the cervix to spermatozoa as well as sperm distribution after natural mating was evaluated by flushing the vagina, the uterus and the uterine tube. For the study of sperm distribution in the female reproductive tract, two methods, flushing and tissue sectioning were evaluated and compared. Using a specially designed catheter it was possible to catheterise the cervix during interoestrus, oestrus, metoestrus and postpartum and to introduce contrast fluids into the uterus to study uterine appearance using hysterography also when the cervix was closed. The period when the cervix was patent was found to vary among individuals: the cervix was open either only during late-oestrus; during midoestrus and late-oestrus; or throughout the entire behavioural oestrus. Patency of the cervix was found to usually coincide with the maximum degree of vaginal cornification and thus, presumably is regulated at least in part by the serum concentration of oestradiol-17β. Hysterograms revealed differences in uterine luminal shape that corresponded to the histological characteristics of the endometrium in cats at various stages of the oestrous cycle, cats given medroxyprogesterone acetate (MPA) and cats with uterine pathology. Straight- and wavy-shaped uterine lumens were characteristic of the uterine horns in the inactive and follicular stages of the oestrous cycle, respectively. Coil-shaped uterine lumen appeared to be a progestagenic effect seen in the luteal, the MPA-treated and the pathological groups. A coiled uterine cavity was suggestive of endometrial hyperplasia, whereas irregular filling defects were indicative of generalised cystic changes in the endometrium. The expression of PCNA in luminal and glandular epithelial cells was observed although the mitotic activity was not related to neither stages of oestrous cycle nor uterine pathological conditions. The results from the sperm distribution study demonstrated that the cervix and the uterotubal junction (UTJ) were sperm barriers in the cat. The endometrial crypts and the UTJ functioned as sperm reservoirs before ovulation, whereas the isthmus was a sperm reservoir around the time of ovulation. The observations determined the dynamics of the cervix, the uterus and the uterine tube in the cat in relation to ovarian activity. This thesis is the first to provide nomenclatures for describing uterine appearance in hysterograms according to the shape of the uterine horns, luminal cavity and the characteristics of the intraluminal lining, to demonstrate the PCNA expression as well as to determine the distribution of spermatozoa in the female reproductive tract of the cat after natural mating

Cell cycle analysis and interspecies nuclear transfer of cat cells treated with chemical inhibitors

This study investigated the effect of chemical inhibitors on the cell-cycle synchronisation in cat fibroblast cells and evaluated the development of interspecies embryos reconstructed from cat donor cells and enucleated bovine oocytes. Cat fibroblast cells were treated with 15 μg/mL roscovitine or 0.05 μg/mL deme-colcine prior to cell cycle analysis and nuclear transfer. The percentage of cat fibroblast cells arrested at the G0/G1 phase in the roscovitine group was similar to that in the control group without any treatment. The percentage of cells arrested at the G2/M phase was significantly higher in the demecolcine group than in the control group. The fusion rate of interspecies couplets was significantly greater in the roscovitine group than in the control group. Most embryos stopped the development at the 2- or 4-cell stage, and none developed into blastocysts. Chemical inhibitor-induced donor cell cycle synchronisation did not overcome developmental arrest in interspecies cloned embryos

HISTONE H3 MODIFICATION OF ISCNT EMBRYOS

This study aimed to determine the acetylation patterns on histone H3K9/18/23 and the dimethylation pattern on histone H3K9 during early embryogenesis among 50 nM Trichostatin A (TSA)-treated iSCNT cat-cow embryos, untreated iSCNT cat-cow embryos (control) and bovine in vitro fertilisation (IVF) embryos, because TSA-treated iSCNT embryos are able to develop into blastocysts. The results show that the acetylation levels of H3K9/18/23 in the TSA-treated iSCNT and bovine IVF embryos were higher than those in the control embryos at almost all of the examined stages (2 h post-fusion / post-insemination (PF/PI), pronuclear (PN), two-cell, four-cell and eight-cell stages). At 6 h PF/PI the acetylation levels on H3K9/23 in the TSA-treated iSCNT and bovine IVF embryos were lower than those in the control, and there was no difference in the acetylation levels of H3K18 among the three groups. The acetylation levels of H3K9/23 increased either in the TSA-treated iSCNT or and bovine IVF embryos increased when those embryos developed to the PN and two-cell stages. The dimethylation level of H3K9 in the TSA-treated iSCNT embryos resembled that of the bovine IVF embryos at all examined stages (2h PF/PI, 6 h PF/PI and PN stages), and these levels were greater than those of the control. This result suggests that treatment of iSCNT embryos with TSA modifies the patterns of histone acetylation and dimethylation at certain lysine residues in a manner that is comparable with that seen in IVF embryos during early embryogenesis

Serum Insulin-like Growth Factor-1 Is a Biomarker of Testosterone Production and Intact Acrosome in Asian Elephants (Elephas maximus)

The objective of this study was to find relationships among serum IGF-1, serum testosterone, seminal plasma IGF-1 concentrations and semen parameters in Asian elephants (Elephas maximus). A total of 17 ejaculates (one to three ejaculates/bull) were collected from seven captive elephant bulls by performing rectal massage. Before each ejaculation, blood samples were obtained for serum IGF-1 and testosterone assays. Subsequently, the semen characteristics of each ejaculate were evaluated. Mean serum IGF-1 concentration of elephant bulls was estimated as 326.3 ± 114.6 ng/mL (median, 286.2 ng/mL; range, 167.4–542.7 ng/mL). An increase in serum IGF-1 concentration was found to correlate with the percentage of spermatozoa with intact acrosomes. In addition, IGF-1 concentration was positively correlated with testosterone level. However, seminal IGF-1 concentrations could not be detected. In conclusion, our findings suggest that serum IGF-1 concentration is likely a biomarker of normal testicular functions, particularly spermatogenesis in elephants. Moreover, this commercial IGF-1 ELISA is eligible for analyzing serum IGF-1 concentration in Asian elephants

Seasonal Effects on Body Condition and Characteristics of the Estrous Cycle in Captive Asian Elephants (<i>Elephas maximus</i>) in Thailand: A Retrospective Study

The aim of this study was to investigate the effects of season on the body condition score (BCS), the characteristics of the estrous cycle (luteal phase [LPL], follicular phase [FPL], estrous cycle [ECL] lengths, and the start of the luteal phase [SLP] and follicular phase [SFP]), and progesterone levels (baseline and peak) of eight captive Asian elephants (Elephas maximus) in Thailand. From 2014 to 2019, blood samples were collected weekly for serum progesterone enzyme immunoassays (EIAs). Estrous cycles (n = 70), including the luteal and follicular phases, and BCS (n = 70) were recorded. Based on the BCS, the LPL, FPL, and ECL were assigned to the following two groups: normal (BCS = 3.0–4.0, n = 38) and overweight (BCS = 4.5–5.0, n = 32). The findings demonstrated that there was no difference in LPL between the groups. However, in the normal group, the ECL was one week longer (14.9 ± 1.7 vs. 13.9 ± 1.7 weeks; p p = 0.06) than in the overweight group. The mean progesterone level during the rainy, hot, and cool seasons was not statistically different. Based on the yearly averaged BCS from three seasons, the baseline and peak levels of progesterone were classified into the normal (n = 16) and overweight (n = 12) groups. Females with a normal BCS tended to exhibit higher progesterone peak levels (p = 0.08). The majority of peaks appeared during the rainy season (53.57%). The BCS was highest during the hot (4.47) and rainy (4.38) seasons, but not during the cool (4.12) season. The LPL, FPL, and ECL were not affected by the season in which the luteal phase occurred. On the other hand, the rainy season had a significant effect on the SFP, resulting in a longer LPL (p p = 0.01); both were the longest during the rainy season. In conclusion, the effects of season on BCS may be related to characteristics of the estrous cycle and peak progesterone levels. Ultimately, these findings provide ground knowledge to assist elephant managers and owners in planning breeding activities using seasonal effects and BCS measurements in tropical climates

Identification of coronavirus disease marker compounds in sweat with comprehensive two dimensional gas chromatography using multiloop splitter-based non-cryogenic artificial trapping modulation system

This study applied comprehensive two-dimensional gas chromatography-mass spectrometry (GC × GCMS–) for analysis of volatile compounds in headspaces of axillary swab samples of RT-PCR or antigen test kit proven COVID-19 positive and negative volunteers in Bangkok, Thailand. The separation was performed using a conventional Deans switch (DS) based heartcut system with an additional splitter system located between the first (1D) column (30 m) and the DS. The splitter consisted of deactivated fused silica (DFS) columns connected into three loops with the progressively doubled perimeters of the downstream loops. An artificial GC × GC result was obtained by applying a periodic heartcut (H/C) event within every artificial modulation period (PAM) of 1.14 min, selectively transferring the analytes onto the second (2D) column (30 m). Chemometric analysis including feature selection was used to identify significantly altered metabolites between the positive and negative groups. Fourteen significant metabolites were tentatively identified, including p-cymene, linalool, and 2,6,11-trimethyldodecane. The marker peak area thresholds were optimized by generating receiver operating characteristic (ROC) curves showing accuracy, sensitivity and selectivity within the ranges of 94–98%, 93–97% and 94–100%, respectively. Based on these results, we hypothesize that SARS-CoV-2 infection disrupts the metabolism of volatile metabolites in sweat, or impacts the microbiome, changing the volatile profile of sweat in infected patients

Exploring enzymatic antioxidant activities in Asian Elephant (Elephas maximus) sperm and seminal plasma

This study explored the relationships between sperm quality and the activities of antioxidant enzymes (superoxide dismutase, SOD, and glutathione peroxidase, GPx) in sperm (S) and seminal plasma (SP) of Asian elephants, as well as the potential protective role of glutamine in semen extenders during chilled storage. In experiment 1, twenty ejaculates from 11 bulls were analyzed for percentages (%) of sperm motility (%MOT), viability (%VIA), and activities of SOD and GPx. A significant negative correlation was found between %VIA and SP-SOD (r = − 0.71), and positive correlations between %MOT and both S-GPx (r = 0.49) and SP-GPx (r = 0.57). Following these findings, glutamine, a precursor of glutathione and semen antioxidant was introduced in Experiment 2. Each ejaculate (in a total of 8 ejaculates obtained from four bulls) was diluted in Tris-fructose- (EYT) and Tris-glucose-egg yolk extender (TCG), respectively, supplemented with 0 (control), 20 and 40 mM glutamine and stored at 4 °C. Sperm parameters including %MOT, %VIA, functional membrane integrity (%sHOST), and DNA integrity (%DNA) were assessed after 24 and 48 h of storage. Results demonstrated that glutamine did not significantly affect %VIA or %DNA. A significant reduction in %MOT was observed at 24 h, but 20 mM glutamine added in extenders reversed this decline at 48 h. A 40 mM addition of glutamine in TCG markedly reduced %sHOST at 24 h. In conclusion, SOD and GPx activities in the sperm and seminal plasma are indicative of sperm quality. During chilling, using a higher level of glutamine may damage the spermatozoa of Asian elephants