Aging related obesity and type 2 diabetes mellitus suppress neuromuscular communication and aggravate skeletal muscle dysfunction in rhesus monkeys.

Age-related functional deterioration in skeletal muscle raises the risk for falls, disability, and mortality in the elderly, particularly in obese people or those with type 2 diabetes mellitus (T2D). However, the response of the skeletal muscle to transitioning from obesity to diabetes remains poorly defined, despite that obesity is classified as a stage of pre-diabetes. We screened and selected spontaneously obese and diabetic rhesus monkeys and examined altered protein expression in skeletal muscle of healthy aging (CON), obesity aging (OB), and type 2 diabetes mellitus aging (T2D) rhesus monkeys using Tandem Mass Tags (TMT)-based quantitative proteomic analysis. In total, we identified 142 differentially expressed proteins. Muscle-nerve communication proteins were firstly suppressed at obese-stage. With the disintegration of skeletal muscle, mitochondrial complex I and other energy homeostasis relate proteins were significantly disordered at T2D stage. Indicating that aging related obesity suppressed muscle-nerve communication and contribute to T2D related functional deterioration of skeletal muscles in elderly rhesus monkeys. Some alterations of muscular functional regulator are detected in both obesity and T2D samples, suggesting some T2D related skeletal muscular hypofunctions are occurring at obesity or pre-obesity stage. Muscle-nerve communication proteins and muscular function related proteins could be potential therapy target or early diagnose marker of for skeletal muscular hypofunctions in aging obesity populations.

Effects of sugar cane extract on steroidogenesis in testicular interstitial cells of male Japanese quail (Coturnix japonica).

Sugar cane extract (SCE) is the end product of glucose, fructose, and sucrose elimination in molasses. SCE has various biological effects, such as anti-inflammation and antioxidation, and it is commonly found in animal feed. The present research is aimed at investigating the reproductive endocrine influence of SCE in male Japanese quails (Coturnix japonica) by feeding SCE containing food. In addition, in vitro Leydig cell culture was conducted to clarify the mechanism of SCE's influence. Our results showed that SCE feed extended the latency to the first neck grab, decreased male quail testis and epididymis weights, cloaca gland size, and reduced serum testosterone concentrations. Steroidogenic enzymes 3ßHSD, 17ßHSD, P450c17, and P450scc gene expression in the testis were decreased in the SCE groups. Western blot analysis showed decreased 3ßHSD in the testis after feeding SCE. Isolated testicular interstitial cells cultured with SCE and ovine-LH suppressed testosterone secretion and 3ßHSD gene expression. In conclusion, SCE as a feed additive has an impact on the sexual behavior and reproductive function of male Japanese quail, with the suppression of steroidogenesis in the Leydig cell. Our results may provide beneficial information to the livestock management and the poultry industry.

Efficient Generation of P53 Biallelic Mutations in Diannan Miniature Pigs Using RNA-Guided Base Editing.

The base editing 3 (BE3) system, a single-base gene editing technology developed using CRISPR/Cas9n, has a broad range of applications for human disease model construction and gene therapy, as it is highly efficient, accurate, and non-destructive. P53 mutations are present in more than 50% of human malignancies. Due to the similarities between humans and pigs at the molecular level, pig models carrying P53 mutations can be used to research the mechanism of tumorigenesis and improve tumor diagnosis and treatment. According to pathogenic mutations of the human P53 gene at W146* and Q100*, sgRNAs were designed to target exon 4 and exon 5 of the porcine P53 gene. The target editing efficiencies of the two sgRNAs were 61.9% and 50.0%, respectively. The editing efficiency of the BE3 system was highest (about 60%) when C (or G) was at the 5th base. Puromycin screening revealed that 75.0% (21/28) and 68.7% (22/32) of cell colonies contained a P53 mutation at sgRNA-Exon5 and sgRNA-Exon4, respectively. The reconstructed embryos from sgRNA-Exon5-5# were transferred into six recipient gilts, all of which aborted. The reconstructed embryos from sgRNA-Exon4-7# were transferred into 6 recipient gilts, 3 of which became pregnant, resulting in 14 live and 3 dead piglets. Sequencing analyses of the target site confirmed 1 P53 monoallelic mutation and 16 biallelic mutations. The qPCR analysis showed that the P53 mRNA expression level was significantly decreased in different tissues of the P53 mutant piglets (p < 0.05). Additionally, confocal microscopy and western blot analysis revealed an absence of P53 expression in the P53 mutant fibroblasts, livers, and lung tissues. In conclusion, a porcine cancer model with a P53 point mutation can be obtained via the BE3 system and somatic cell nuclear transfer (SCNT).

Ovarian progesterone suppresses depression and anxiety-like behaviors by increasing the Lactobacillus population of gut microbiota in ovariectomized mice.

Depression and anxiety, which are severe symptoms during menopause, are caused by ceased ovarian activity and declined serum progesterone levels. Studies have demonstrated that gut microbiota can regulate brain function and change the microbiota composition during the perimenopause period. This study investigated whether progesterone affects depressant and anxious behaviors via gut microbiota. In ovariectomized (OVX) mice, treatment with progesterone improved depressive and anxious behaviors, and gut microbiota composition was significantly changed. In particular, increased Lactobacillus spp. were observed in these mice. Reduction of microbiota by antibiotic treatment abolished the effect of progesterone on depression and anxiety. In addition, administration of Lactobacillus (L.) reuteri that was increased by progesterone also reduced the depressant behavior in OVX mice, and BDNF gene expression was elevated by progesterone treatment and L. reuteri administration in the hippocampus. Moreover, we found that progesterone stimulated the growth of L. reuteri in vitro. In summary, our findings indicate that progesterone reduces depression and anxiety through changes in gut microbiota composition, particularly by increasing the Lactobacillus spp. population.

The relation between liver damage and reproduction in female Japanese quail (Coturnix japonica) exposed to high ambient temperature.

Reproductive failure associated with heat stress is a well-known phenomenon in poultry. High temperatures also induce various metabolic disturbances in many animals. Because the liver plays a central role in metabolism, the present study aimed to clarify the relationship between liver and reproduction in Japanese quails exposed to high temperatures. In the consecutive 20-D experimental period, quails were treated with 25°C (control) or 34°C (heat) from 12:00 to 16:00. Eggs were collected for hatching. On completion of the experimental period, quails were humanely euthanized for hormone analyses (e.g., serum and ovarian follicles). Serum metabolites were analyzed using GC/MS. Liver and ovary samples were collected for mRNA levels, histomorphology, and metabolic analysis. Ovary and oviduct weights significantly decreased after daily heat exposure. The number and weight of hierarchical follicles also decreased. Consequently, egg weight decreased. Although there was no difference in fertilization rate, chick birth weight significantly decreased in the heated group. Corticosterone and 17ß-estradiol in the serum significantly increased in the heated group. Yolk corticosterone and 17ß-estradiol concentration and content were higher in the heated group. Ovary sterologenic enzymes gene P450scc and estrogen receptor expression level increased. The FSH receptor decreased in heat-stressed quails. MetaboAnalyst analysis indicated that high temperature affects propanoate metabolism, beta-alanine metabolism, aspartate metabolism, and histidine metabolism. Triglyceride and cholesterol levels in the liver increased in the heated group. The heated group also had an increased mRNA expression of AGPAT5, apoptosis gene caspase3, and the immunocytokine genes IL-6 and TLR4. However, NF-κB gene expression decreased. These results suggest that high temperatures affect lipid metabolism and apoptosis and cause inflammation in the liver. High temperature induced ovarian dysfunction, which resulted in the decline of hierarchical follicle number and weight, egg weight, and chick birth weight. The increased level of 17ß-estradiol suggests liver damage. Protecting liver function from damage may assist quails cope in summer.

Yolk immunoreactive corticosterone in hierarchical follicles of Japanese quail (Coturnix japonica) exposed to heat challenge.

High temperature decreases the egg number, ovarian weight, and hierarchical follicle number. In the present study, we investigated the effect of high temperature on the quality of eggs of adult female quails. Laying quail were raised under a standard thermal condition of 25 °C until exposed to an elevated temperature of 34 °C (experimental) or maintained at 25 °C (control) from 12:00 to 16:00 for 10 consecutive days. Weight and number of eggs were measured; serum and the largest follicles were collected and used for hormone measurement. Ovaries and adrenals were collected for expression analysis of 3ß- and 17ß-HSD, genes encoding steroidogenic enzymes. Egg weight slightly decreased with an increase in the treatment time in the heat-challenged group; the egg weight significantly decreased in the heat treatment group than in the control group during the last 2 days of experiment (P < 0.05). The laying rate showed no difference during the experimental period but significantly decreased on the last day in the heat treatment group. In the experimental group the ovaries and oviducts were lighter (P < 0.05) and the hierarchical follicle number and ovarian weight decreased (P < 0.05) compared to the control group. Although serum corticosterone level significantly increased after heat challenge (P < 0.05) and immediately recovered to the normal level, yolk immunoreactive corticosterone in the hierarchical follicle (F1, F2, F3) significantly increased (P < 0.05). The expression level of 17ß-HSD showed no changes in the ovary but significantly increased in adrenals (P < 0.05). Our findings indicate that the effects of heat challenge on the maternal ovary in the quail are mediated through the adrenal function.

Heat challenge influences serum metabolites concentrations and liver lipid metabolism in Japanese quail (Coturnix japonica).

High temperature induces various metabolic disturbances in animals. However, no comprehensive information is currently available on the metabolic pathway affected by high environmental temperature. The present study examined metabolite content in the serum of heat challenged quails using metabolomic analysis. In the present study, female quails with normal laying rate at 20 weeks kept in standard condition (control group) or exposed to 34°C 4 hr per day (12:00 to 16:00 hr)(heat group) for 10 consecutive days. The metabolomic analysis identified 165 metabolites in the serum, and significant differences were observed in the serum for 7 metabolites between two groups. An analysis by MetaboAnalyst, a web-based metabolome data tool, indicate that high temperature affect ketone body metabolism, butyrate metabolism, arginine and proline metabolism. Furthermore, histological examination of liver indicates a heat challenge induced abnormal lipid metabolism. Triglyceride and cholesterol level in the liver increased, however cholesterol level decreased in the serum. Genes related to lipid metabolism significantly increased in the liver after heat challenge. The present study demonstrated that high temperature cause liver damage, thus lipid metabolic was affected. Protect liver under high temperature could be one solution for coping with high temperatures in summer.

Male Hatano low-avoidance rats show more active sexual behavior with lower plasma testosterone than high-avoidance rats.

Two inbred strains of Sprague-Dawley rats, known as the Hatano high- and low-avoidance animals (HAA and LAA respectively), have been selectively bred for high versus low rates of avoidance responses in a shuttle-box avoidance task. To investigate differences in the sexual behavior of Hatano rats, male HAA, LAA and SD rats were tested from 12 to 15 weeks of age. LAA rats exhibited more rapid and frequent sexual behavior than HAA or SD rats, and such differences increased with repeated sexual experience. Plasma testosterone levels tended to be lower in LAA rats than in HAA or SD rats, suggesting that active sexual behavior in LAA rats is not related to these levels. Strain differences in mating behavior between HAA and LAA rats may be caused by emotional responses to novelty.

Supplemental dietary phytosterin protects against 4-nitrophenol-induced oxidative stress and apoptosis in rat testes.

4-Nitrophenol (PNP), is generally regarded as an environmental endocrine disruptor (EED). Phytosterin (PS), a new feed additive, possesses highly efficient antioxidant activities. The transcription factor, nuclear factor-erythroid 2-related factor 2 (Nrf2), is an important regulator of cellular oxidative stress. Using rats, this study examined PNP-induced testicular oxidative damage and PS-mediated protection against that damage. The generation of MDA and H2O2 upon PNP and PS treatment was milder than that upon treatment with PNP alone. This mitigation was accompanied by partially reversed changes in SOD, CAT, GSH and GSH-Px. Moreover, PNP significantly reduced the caudal epididymal sperm counts and serum testosterone levels. Typical morphological changes were also observed in the testes of PNP-treated animals. PNP reduced the transcriptional level of Nrf2, as evaluated by RT-PCR, but it promoted the dissociation from the Nrf2 complex, stabilization and translocation into the nucleus, as evaluated by immunohistochemistry and Western blotting. In addition, PNP enhanced the Nrf2-dependent gene expression of heme oxygenase-1 (HO-1) and glutamate-cysteine ligase catalytic subunit (GCLC). These results suggest that the Nrf2 pathway plays an important role in PNP-induced oxidative damage and that PS possesses modulatory effects on PNP-induced oxidative damage in rat testes.

Tert-butylhydroquinone attenuates scrotal heat-induced damage by regulating Nrf2-antioxidant system in the mouse testis.

Tert-butylhydroquinone (tBHQ), a widely used nuclear factor erythroid 2-related factor 2 (Nrf2) activator, was always employed to investigate the potential protective role of Nrf2 activation. In this study, to elucidate the effect of tBHQ on scrotal heat-induced damage and Nrf2-antioxidant system in mouse testes, eight-week-old mice were administrated with or without dietary tBHQ (1%w/w) for 1week and afterward subjected to a single scrotal heat treatment (42°C for 25min). Trunk blood and testes were collected 3h or 1, 2, or 7days later. Mice displayed less germ cell loss in testes, higher relative testis weight and lower testosterone concentration on day 2 in tBHQ treatment group. Before heat treatment, there were significant increases in malondialdehyde (MDA) concentration in tBHQ treatment group. After heat treatment, mice in tBHQ treatment group showed lower MDA concentration than those in non-tBHQ treatment group. In addition, both tBHQ pretreatment and scrotal heat treatment induced markedly increased Nrf2 protein expression in cytoplasm and nuclei of interstitial cells, accompanying with elevated mRNA expression of Nrf2 and Nrf2-regulated genes in mice testes. Our data indicated that pretreatment to tBHQ induced a mild oxidative stress, and further enhanced the cellular antioxidative ability to protect testicular cells against scrotal heat-induced damage via a mechanism that might involve the Nrf2-antioxidant system in mice testes.