Proteomic analysis of ITPR2 as a new therapeutic target for curcumin protection against AFB1-induced pyroptosis.

Aflatoxin B1 (AFB1) is extremely carcinogenic and can cause liver cancer in humans and animals with continued ingestion. As a natural compound, curcumin (Cur) exhibits excellent anti-inflammatory, and anti-cancer properties with few side effects. In this study, a total of 60 male mice (6-week-olds, 15 per group). After one week of acclimatization feeding, the mice were divided into control group (Con), AFB1 group, curcumin group (Cur), and AF+Cur group. The mice were gavaged with curcumin (Cur, 100 mg/kg) and/or AFB1 (0.75 mg/kg). To identify a new therapeutic target for AFB1-induced pyroptosis, we performed proteomic profiling for curcumin alleviating liver injury caused by AFB1 to further validate the targets through volcano plot analysis, Venn analysis, heatmap analysis, correlation, cluster analysis, GO and KEGG enrichment. AFB1 exposure resulted in the loss of hepatocyte membrane, swelling of the endoplasmic reticulum, and a significant increase in transaminase (ALT and AST) contents, while curcumin greatly improved these changes. We found that differentially expressed proteins are enriched in the endoplasmic reticulum membrane and identified ITPR2 as a target of curcumin that alleviates AFB1-induced liver injury by proteomics. Furthermore, ITPR2 expression was detected by immunofluorescence, and qRT-PCR for mRNA expression of genes downstream of ITPR2 (calpain1, calpain2, caspase-12, caspase-3). ITPR2-activated endoplasmic reticulum stress-related proteins (calpain1, calpaini2, bcl-2, BAX, cl-caspase-12, cl-caspase-3), apoptosis (PARP) and pyroptosis (DFNA5) related proteins were examined by western blotting. The analysis showed that it effectively prevents AFB1-induced pyroptosis by lowering endoplasmic reticulum stress via interfering with ITPR2 and its downstream proteins (calpain1, calpain2, bcl-2, Bax) and inhibiting caspase-12/caspase-3 pathway. Conclusively, this study applied proteomic profiling to elucidate ITPR2 as a new target, which might give a new perspective on the mechanism of curcumin alleviating AFB1-induced pyroptosis.

Effect of Curcumin as Feed Supplement on Immune Response and Pathological Changes of Broilers Exposed to Aflatoxin B1.

In this study, we examined the protective effects of curcumin against the AFB1-induced immune response of and pathological changes in broilers. Histopathology examinations showed that at day 28, AFB1 (5 mg/kg) exposure leads to severe histological changes in the spleen, thymus and bursa of Fabricius with a decrease in the number and karyoplasmic area ratio of plasma cells. Curcumin alleviated the AFB1-induced immune organs' damage as well as the changes in plasma cells in a dose-dependent manner. RT-PCR data showed that AFB1 significantly downregulated the IL-2 and IFN-γ mRNA expression levels in the thymus, spleen and bursa of Fabricius. However, curcumin supplementation improved the AFB1-induced immune organs' damage via upregulated cytokines' expression. Intriguingly, similar trends were noticed in abnormal morphological changes and the immune response at day 35 after the withdrawal of AFB1 and curcumin from the diet, suggesting the protective effects and immunomodulatory function against AFB1 in broilers. The current study provides a scientific experimental basis for the application of curcumin as a therapeutic drug or additive in animal husbandry productive practice.

Melatonin protects against maternal diabetes-associated meiotic defects by maintaining mitochondrial function.

Maternal diabetes has been widely reported to adversely affect oocyte quality. Although various molecules and pathways may be involved in this process, strategies to prevent maternal diabetes-induced deterioration of oocyte quality remain unexplored. Melatonin is synthesized by the pineal gland and has been shown to have beneficial effects on oocyte quality owing to its antioxidative function. In the present study, we found that the exposure of oocytes of diabetic mice to melatonin, in vitro, alleviated aberrant oocyte maturation competence. Notably, melatonin supplementation attenuated defects in spindle organization and chromosome alignment by mediating the expression of TPX2 and pericentrin localization. Importantly, melatonin eliminated the accumulation of reactive oxygen species and increased the cytosolic Ca2+ levels in diabetic oocytes by maintaining mitochondrial function. Moreover, the occurrence of autophagy and apoptosis was reversed in diabetic oocytes after melatonin exposure via decreased LC3ß expression. Collectively, our findings provide evidence that melatonin supplementation can protect oocytes from maternal diabetes-related meiotic defects and poor egg quality, providing a potential strategy for improving oocyte quality in assisted reproductive technologies.

The synergistic effects of applying low-level laser therapy plus ultrasound on pain and muscle function in patients with knee osteoarthritis: A protocol of a randomized double-blind study.

BACKGROUND: To our knowledge, only 1 study with limited sample size tried to evaluate the synergistic effects of ultrasound and low-level laser therapy (LLLT) in patients with knee osteoarthritis. Further research is needed to confirm this synergy with larger numbers and better design. Therefore, we will conduct this present randomized double-blind study to evaluate the synergistic effects of simultaneously applying ultrasound plus LLLT on pain and muscle function in patients with knee osteoarthritis. METHODS: The study protocol is a randomized, controlled, double-blind design. The study will be conducted at our academic hospital from February 2021 to January 2022. The study protocol was approved through Institutional Review Board in the Hunan Provincial People's Hospital. Patients will be assigned at random to the ultrasound + LLLT group, LLLT group, or the ultrasound group. After baseline examination, all patients will be given a full explanation of the treatment protocol and will be required to sign a written informed consent for study participation and for publication of the results. All the data collectors, surgeons, statistical analysts, as well as result assessors are not aware of grouping assignment. The primary outcome is weekly change in pain intensity relative to baseline through 6 weeks of therapy. RESULTS: This protocol will provide a reliable theoretical basis for the following research. CONCLUSION: It is assumed that there will be a remarkable difference in postoperative outcomes between the intervention and control groups. TRIAL REGISTRATION: This study protocol was registered in Research Registry (researchregistry6470).

Curcumin ameliorates duodenal toxicity of AFB1 in chicken through inducing P-glycoprotein and downregulating cytochrome P450 enzymes.

It has been reported that oral intake of aflatoxin B1 (AFB1)-contaminated feed could cause acute, sub-chronic, or chronic toxicity in livestock and poultry. However, the harmful effect of AFB1 on the small intestine is still controversial. Therefore, blocking the entry of AFB1 into the body through the digestive tract is one of the important methods to prevent its toxicity. In the present study, 1-day-old Arbor Acres broilers were randomly divided into 6 groups including control group, curcumin control group (450 mg curcumin/kg feed), curcumin low-, medium-, and high-dose group (150, 300, and 450 mg curcumin/kg feed + 5 mg AFB1/kg feed), and AFB1 group (5 mg AFB1/kg feed). After 28 d, the samples of chickens' duodenums were collected for further analyses. AFB1 caused abnormal functional and morphological changes in the duodenum, including histological lesions, increased the length of the duodenum and depth of crypt, decreased the unit weight of the duodenum, height of villus, and the value of villus height/crypt depth. Meanwhile, AFB1 administration enhanced malonaldehyde activity, 8-HOdG level, and the mRNA expression of cytochrome P450 (CYP450) enzymes, and reduced superoxide dismutase, catalase, adenosine triphosphatase (ATPase) activity and the mRNA expression of Abcb1. Importantly, curcumin supplementation partially ameliorated AFB1-induced abnormal functional and morphological signs of the duodenum, alleviated AFB1-induced oxidative stress, and decreased the mRNA expression of CYP450 enzymes. Furthermore, curcumin ameliorated AFB1-induced decrease in the Abcb1 mRNA expression, P-glycoprotein (P-gp) level, and ATPase activities. It has been suggested from these results that curcumin supplementation in the feed could ameliorate AFB1-induced duodenal toxicity and damage through downregulating CYP450 enzymes, promoting ATPase activities, and inducing P-gp in chickens.

Detection of Aflatoxin adducts as potential markers and the role of curcumin in alleviating AFB1-induced liver damage in chickens.

In this study, we identified AFB1 adducts as potential markers and investigated the role of curcumin in alleviating AFB1-induced liver damage by suppressing the production of AFB1 adducts and oxidative stress in AA broilers liver. A total of 64 one-day-old Arbor Acres (AA) broilers were randomly divided into four groups, including control group, AFB1 group (5 mg/kg AFB1), cur + AFB1 group (300 mg/kg curcumin+5 mg/kg AFB1) and curcumin group (300 mg/kg). Serum biochemical parameters, liver antioxidant abilities, AFB1 adducts and oxidative stress mechanism were studied in broilers. AFB1 administration accompany with signs of liver injury, including hepatic histological lesions, increased serum enzymes activities, decreased liver antioxidant enzymes activities and the suppression of ROS and 8-OHdG. Meanwhile, Nrf2/HO-1 pathway was depressed by AFB1 treatment. Immunohistochemistry and ELISA showed that AFB1 significantly increased AFB1-DNA adduct in liver (p < 0.05) and AFB1-lysine adduct in serum (p < 0.05). Importantly, supplementation of curcumin can ameliorate these alterations. Intriguingly, curcumin alleviated AFB1-induced toxicity and oxidative stress by inhibiting the generation of ROS, 8-OHdG and AFB1 adducts, and activated Nrf2 signaling pathway in broilers. Conclusively, our experiments suggest that curcumin could be considered as a potential agent for prevention of AFB1-induced toxicity and oxidative stress, and AFB1 adducts could be suitable therapeutic targets.