Single-cell RNA sequencing reveals the differentiation and regulation of endplate cells in human intervertebral disc degeneration.

Low back pain (LBP) is largely attributed to intervertebral disc degeneration (IVDD), of which the endplate changes are an important component. However, the alterations in cell fate and properties within the endplates during degeneration remain unknown. Here, we firstly performed the single-cell RNA-sequencing analysis (scRNA-seq) of the cells focusing on degenerative human endplates. By unsupervised clustering of the 8,534 single-cell based on the gene expression, we identified nine distinct cell types. We employed Gene ontology (GO) analysis, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways analysis, and the single-cell regulatory network inference and clustering (SCENIC) to determine the enriched pathways and transcriptional activities across seven chondrocyte subpopulations. Furthermore, two cell fates of chondrocyte differentiation were found by trajectory analysis, one was enriched in inflammation-related genes, and the other was related to extracellular matrix (ECM). Additionally, the intercellular interactions of macrophages (MA) and chondrocytes, T cells/natural killer cells (T/NK) and chondrocytes were examined by ligand-receptor pairs analysis, showing the important regulative function of FN1 from MA and CD74 from T/NK during endplate degeneration. Overall, our findings provide novel perspectives on the endplate degeneration at the single-cell level and a whole-transcriptome size.

METTL3 Promotes Nucleus Pulposus Cell Senescence in Intervertebral Disc Degeneration by Regulating TLR2 m6A Methylation and Gut Microbiota.

BACKGROUND: Nucleus pulposus cell (NPC) senescence in intervertebral disc (IVD) tissue is the major pathological cause of intervertebral disc degeneration (IDD). N6-methyladenosine (m6A) methylation and gut microbiota play important roles in the progression of IDD. This study investigated whether methyltransferase-like 3 (METTL3) regulates TLR2 m6A modification and gut microbiota to influence NPC senescence. METHODS: An IDD rat model was established by lumbar IVD puncture and NPCs were challenged with IL-1ß to mimic IVD injury. IDD rats and IL-1ß-exposed NPCs were treated with METTL3-interfering lentivirus and the TLR2 agonist Pam3CSK4. Compositional changes in the rat gut microbiota were analyzed and fecal microbiota transplantation procedures were used. NPC senescence, cell cycle, and the expression of senescence-associated secretory phenotype (SASP) factors were assessed. The m6A enrichment of TLR2 and the binding of IGF2BP1 to TLR2 mRNA were examined. RESULTS: METTL3 and TLR2 were highly expressed in IDD rats. METTL3 silencing attenuated senescent phenotypes and reduced secretion of SASP factors. Pam3CSK4 reversed the beneficial effects of METTL3 silencing on NPC senescence and IVD injury. METTL3 stabilized TLR2 mRNA in an IGF2BP1-dependent manner. METTL3 silencing restored specific gut microbiota levels in IDD rats, which was further reversed by administration of Pam3CSK4. Fecal microbiota from METTL3 silenced IDD rats altered the pathological phenotypes of IDD rats. CONCLUSIONS: These results demonstrate the beneficial effects of METTL3 silencing on NPC senescence and amelioration of IVD injury, involving modulation of TLR2 m6A modification and gut microbiota. These findings support METTL3 silencing as a potential therapeutic target for IDD.

Senescent-like macrophages mediate angiogenesis for endplate sclerosis via IL-10 secretion in male mice.

Endplate sclerosis is a notable aspect of spine degeneration or aging, but the mechanisms remain unclear. Here, we report that senescent macrophages accumulate in the sclerotic endplates of lumbar spine instability (LSI) or aging male mouse model. Specifically, knockout of cdkn2a (p16) in macrophages abrogates LSI or aging-induced angiogenesis and sclerosis in the endplates. Furthermore, both in vivo and in vitro studies indicate that IL-10 is the primary elevated cytokine of senescence-related secretory phenotype (SASP). Mechanistically, IL-10 increases pSTAT3 in endothelial cells, leading to pSTAT3 directly binding to the promoters of Vegfa, Mmp2, and Pdgfb to encourage their production, resulting in angiogenesis. This study provides information on understanding the link between immune senescence and endplate sclerosis, which might be useful for therapeutic approaches.

Evaluating the long-term cost-effectiveness of fixed-ratio combination insulin degludec/liraglutide (IDegLira) versus other treatment regimens in the chinese type 2 diabetes patients.

BACKGROUND AND AIMS: To assess the cost-effectiveness of utilizing IDegLira in comparison to other treatment regimens ( liraglutide and degludec) in managing type 2 diabetes, taking into account the Chinese healthcare system's perspective. METHODS: The clinical data were obtained from the randomized controlled trials (RCTs) of the DUAL I and DUAL II evidence studies that took place in China. To estimate the lifetime quality-adjusted life-years (QALYs) and direct medical costs of patients receiving different treatment strategies from a long-term perspective, the IQVIA CORE Diabetes Model version 9.0 (IQVIA, Basel, Switzerland) was utilized. The costs were evaluated from the perspective of the China National Health System. Future costs and clinical benefits were discounted annually at 5%, and sensitivity analyses were conducted. RESULTS: IDegLira was projected to reduce the incidence of diabetes-related complications and improve quality-adjusted life expectancy (QALE) versus liraglutide and degludec. A survival benefit was observed with IDegLira over Liraglutide (0.073 years). Lifetime costs were lower by Chinese yuan (CNY) 27,945 on IDegLira than on Liraglutide therapy. A similar survival benefit was observed with IDegLira over degludec (0.068 years). Lifetime costs were lower by CNY 1196 on IDegLira than on degludec therapy. Therefore, IDegLira was found to be cost-effective versus liraglutide and degludec with incremental cost-effectiveness ratios of Dominant per QALY gained, respectively, under the threshold of three times the gross domestic product (GDP) per capita in China. CONCLUSION: IDegLira is a cost-effective hypoglycemic treatment option that delivers positive clinical outcomes while also reducing costs for Chinese patients living with type 2 diabetes.

Production performance and rumen bacterial community structure of Hu sheep fed fermented spent mushroom substrate from Pleurotus eryngii.

This study aimed to investigate the effect of fermented spent mushroom substrate from Pleurotus eryngii (SMPE) supplementation on production performance, meat quality and rumen bacterial community structure of Hu sheep. 120 2-month-old Hu sheep with average body weight [(13.50 ± 3.10) kg] were selected and randomly divided into 4 groups with 3 replicates per group and 10 sheep per replicate. The control group (RL1) was fed a total mixed ration (TMR), and group RL2, RL3 and RL4 were fed the basal diets supplemented with 15%, 30% and 45% fermented SMPE, respectively. The pretest period lasted for 10 days and the test period lasted for 150 days. The results showed that: (1) Difference (p < 0.05) was observed in average daily feed intake (ADFI) and feed conversion ratio (FCR) between RL2 and RL4 groups. The eye muscle area (EMA) and grade rule (GR) values in RL2 and RL3 were significantly higher than those in RL1 and RL4 groups (p < 0.05). (2) The contents of threonine, valerine, leucine, lysine, histidine, essential amino acids, flavor amino acids, aspartic acid, serine, glutamic acid and arginine of the longissimus dorsi muscle in RL2 and RL3 groups were significantly higher than RL1 and RL4 (p < 0.05). (3) A total of 1,202,445 valid sequences were obtained from rumen of Hu sheep fed different amounts of fermented feed, and the valid sequences were clustered into 9824 Operational Taxonomic Units (OTUs). (4) α diversity analysis showed that the richness and diversity of rumen bacterial communities in Hu sheep in RL1, RL2, RL3 and RL4 groups were significantly higher than RL0 (raw materials of fermented SMPE) group (p < 0.05). ß diversity analysis showed that the bacterial community structure was the most different between RL0 and RL3. (5) At the genus level, compared with RL1, the relative abundance of Christensenellaceae R-7 in RL3 group decreased significantly by 33.59%, the relative abundance of Prevotellaceae UCG001 in RL2, RL3 and RL4 decreased significantly by 50.41%, 62.24% and 49.17%, respectively, and the relative abundance of Ruminococcaceae NK4A214 in RL2 group increased significantly by 35.01% (p < 0.05). In summary, the addition of fermented SMPE to TMR can significantly improve the production performance, meat quality and rumen bacterial community diversity and abundance of Hu sheep.

Chinese herb feed additives improved the growth performance, meat quality, and nutrient digestibility parameters of pigs.

BACKGROUND: Since the use of antibiotics in animal feed has become a critical concern worldwide due to severe threats to human health and environment, we are in need of finding alternatives to antibiotics in pig breeding, maintaining the health of pigs, and getting high-quality pork. As traditional Chinese herbs (TCH) are rich natural resources in China and show great benefits to human health we propose to transfer this abundant resource into animal production industry as additives. METHODS: Three groups of Chinese herbs (groups A, B, and C) were used as feed additives in the diet for pigs. In total 32 pigs were arranged in four groups (groups A, B, C, and control group, NC), fed in the same facility, eight pigs (one group) in each colony, free drinking, for 120 days. The feed:gain ratio (F/G), meat quality, total protein, and amino acid concentration of muscle were checked in the experiments. RESULTS: After 120 days of feeding, the feed:gain ratio (F/G) of pigs in groups A, B, and C was decreased 17.56%, 9.31%, and 13.86% compared with NC treatment, respectively. The diets supplemented with Chinese herbs improved meat quality, increased loin eye area (especially group A and C showed significant difference, P < .001), the total protein (increased ratio vs NC was A = 4.54%, B = 0.38% and C = 3.53%), amino acid concentration of muscle, increased the villus height:crypt depth ratio, and induced positive effects on serum biochemical parameters and immune function (serum TC and TG concentrations were significantly lower than those in the NC group, P < .05.). CONCLUSIONS: The use of Chinese herbal feed additives can reduce the cost of pig breeding and produce high-quality pock. The combination of these effects would contribute to better absorption ability of the intestinal tract and yield a better growth performance.