We present an active-matrix digital microfluidics platform for bio-sample handling, which demonstrates extension of the ubiquitous and highly-scalable flat-panel technology from displays to bio-applications, and leverage the well-established a-Si thin-film transistor (TFT) manufacturing facilities and excess capacity. The fabricated chip contains 32 × 32 pixels in an active area of 10 cm ² , which can be individually or simultaneously addressed. Generation and manipulation of liquid droplets at single pixel level with the volume coefficient of variation around 1% is demonstrated. Such performance is substantial improvement over current state-of-the-art (9-pixel resolution and 4% variation). This work opens new possibilities for on-chip in vitro diagnostics in point-of-care testing.

Heat stress is one of the major stressors affecting dairy cow production, which causes great losses in animal husbandry production and economic development. This study combined 16S rDNA sequencing, metagenomic sequencing, and metabolomic analysis to investigate the effects of heat stress on the rumen microbiome and metabolism of cows. To achieve this, 10 cows each of growing heifers, heifers, and lactating cows were selected for sample collection in April and August (a total of 60 cows). Ruminal fluid was collected, filtered through gauze, and immediately transferred to liquid nitrogen prior to metagenomic, 16S rDNA sequencing, and metabolomic analyses. Heat stress increased the abundance of pathogenic bacteria, such as Treponema_2 , Sphingobacterium , and Streptococcus , in cattle at all growth stages, with the structure and composition of rumen microbial communities being the most affected by heat stress in growing heifers and the least affected by heat stress in lactating cows. Heat stress led to a significant decrease in carbohydrate metabolic function in growing heifers and heifers, whereas microbial function was the most stable in lactating cows, suggesting that changes in microbial function are not only more conserved than microbial composition but are also affected by age. Heat stress led to significant enrichment of fatty acyl metabolites in growing heifers, heifers, and lactating cows, and the heat stress indicator species Bacillus , Lactococcus , Streptococcus , and Enterococcus were significantly associated with fatty acyl compounds in growing heifers and heifers. Therefore, heat stress alters the metabolic status of cows by modulating fatty acid biosynthesis, the PPAR signaling pathway, and arachidonic acid metabolism through the regulation of long-chain fatty acids and other biomarkers in cows at three growth stages. IMPORTANCE: Heat stress is one of the main causes of economic losses in the dairy industry worldwide; however, the mechanisms associated with the metabolic and microbial changes in heat stress remain unclear. Here, we characterized both the changes in metabolites, rumen microbial communities, and their functional potential indices derived from rumen fluid and serum samples from cows at different growth stages and under different climates. This study highlights that the rumen microbe may be involved in the regulation of lipid metabolism by modulating the fatty acyl metabolites. Under heat stress, the changes in the metabolic status of growing heifers, heifers, and lactating cows were closely related to arachidonic acid metabolism, fatty acid biosynthesis, and energy metabolism. Moreover, this study provides new markers for further research to understand the effects of heat stress on the physiological metabolism of Holstein cows and the time-dependent changes associated with growth stages.

The objective of this study was to assess the effect of heat stress on the autophagy and apoptosis of the rumen, abomasum, duodenum, liver and kidney in calves. Two groups of Holstein male calves were selected with similar birth weights and health conditions. Heat stress (HT): Six calves (birth weight 42.2 ± 2.3) were raised from July 15 to August 19. Cooling (CL): Six calves (birth weight 41.5 ± 3.1 kg) were raised from April 10 to May 15. All the calves were euthanized following captive bolt gun stunning at 35 d of age. The expression of protein 1 light chain 3-Ⅱ (LC3-Ⅱ) and caspase3 in the rumen, abomasum, duodenum, liver and kidney were determined by western blotting. In addition, other possible relevant serum biochemical parameters were evaluated. Significant differences were observed in alkaline phosphatase (ALP), albumin (ALB) and glucose (Glu). The results showed that heat stress could increase the autophagy and apoptosis of the kidney, duodenum and abomasum. However, heat stress had no effect on the autophagy and apoptosis of the liver. Additionally, the expression of caspase-3 in the rumen in HT was significantly lower than that in CL. In conclusion, the effects of heat stress on autophagy and apoptosis are organ-specific. The results provide knowledge regarding autophagy and autophagy in calf heat stress management.

Background: Long-term high-fat diet (HFD) consumption induces hepatic lipid accumulation and metabolic dysregulation, but its integrated effects on the hepatic proteome and phosphoproteome remain incompletely understood. This study investigated HFD-induced hepatic molecular alterations using quantitative proteomics and phosphoproteomics. Methods: Eight-week-old male C57BL/6J mice were fed a control diet (CON) or HFD for 16 weeks, followed by phenotypic assessment, serum redox assays, and hepatic proteomic and phosphoproteomic analyses. Results: HFD-fed mice exhibited increased body weight, fat mass, liver weight, and marked hepatic lipid accumulation. Blood glucose levels and total pyruvate tolerance test area under the curve (AUC) were increased, whereas baseline-adjusted incremental AUC did not differ significantly. Serum malondialdehyde levels were elevated, whereas glutathione peroxidase activity was reduced, indicating impaired glucose homeostasis and redox balance. Proteomic analysis identified 15 differentially expressed proteins (4 upregulated and 11 downregulated), associated with lipid metabolism, biological oxidation, and xenobiotic metabolism. Gene set enrichment analysis identified significant positive enrichment of Class I peroxisomal membrane protein import and significant negative enrichment of cholesterol biosynthesis, gluconeogenesis, and xenobiotics. Phosphoproteomic analysis identified 8593 phosphoproteins and 28,180 phosphorylation sites, including 2045 differentially phosphorylated sites (1044 increased and 1001 decreased). Enrichment and exploratory network analyses implicated mitophagy, glucose metabolism, mTOR signaling, autophagy, oxidative phosphorylation, and cytoskeletal organization. Conclusions: Long-term HFD feeding induced obesity, glucose dysregulation, hepatic steatosis, and redox imbalance. Fatty acid oxidation and peroxisomal abundance patterns may reflect metabolic adaptation to lipid loading, while phosphosite abundance changes link nutrient sensing and autophagy regulation to the hepatic response.

Heat stress, particularly during the prepartum period, compromises dairy cow health and has lasting effects on offspring. This study aimed to investigate the effects of maternal yeast culture (YC) supplementation during heat stress on the intestinal health of newborn calves, with a specific focus on elucidating the associated changes in jejunal morphology, microbial community, and metabolic profiles. Cows receiving YC had lower rectal temperatures, confirming mitigated heat stress. Their calves showed enhanced immune status (higher IgA, IgG) and reduced oxidative stress and metabolic markers. Jejunal structure and barrier integrity were improved. While microbial diversity was similar, specific bacterial abundances differed (e.g., Enterococcus spp. enriched in YC calves). Metabolomic analysis revealed modulated bile acid pathways and metabolites in YC calves, correlating with the microbial shifts. In conclusion, maternal YC supplementation during heat stress supports calf early-life gut health and development through maternal-offspring programming, optimizing gut microbiota, metabolism, and immunity.

Background: Branched-chain amino acid (BCAA) is involved in glucose and lipid metabolism and energy homeostasis, but whether its effects on maternal metabolism and offspring growth during lactation depend on dietary protein level remains unclear. Methods: C57BL/6J lactating dams were randomized after delivery to a normal-protein control group (CN, 20% protein), low-protein group (LP, 10% protein), CN + BCAA group (CNB), or LP + BCAA group (LPB) (n = 11/group). Maternal body composition, glucose and lipid metabolism, hepatic energy status, oxidative stress indices, and pup litter weight gain were assessed. Results: The LP diet reduced maternal body weight, pup litter weight gain, and glucose responses during the intraperitoneal pyruvate tolerance test (IPPTT), and increased hepatic lipid deposition and oxidative stress. BCAA supplementation increased pup litter weight gain under normal-protein conditions, but decreased maternal food intake and offspring growth under low-protein conditions. Metabolically, BCAA supplementation reduced the homeostasis model assessment of insulin resistance (HOMA-IR) and hepatic PEPCK1 activity under normal-protein conditions, while partially restoring IPPTT glucose responses and increasing hepatic PEPCK1 activity under low-protein conditions. In addition, BCAA supplementation reduced hepatic lipid deposition and ATP content at both protein levels. Notably, under low-protein conditions, BCAA supplementation increased serum β-hydroxybutyrate (β-HB), while alleviating hepatic oxidative stress. Conclusions: BCAA supplementation exerted opposite effects on offspring growth during lactation under different maternal dietary protein levels. These effects may be related to differential changes in maternal glucose and lipid metabolism and hepatic energy status, suggesting that BCAA supplementation during lactation should be evaluated according to maternal dietary protein level.