HBB contributes to individualized aconitine-induced cardiotoxicity in mice via interfering with ABHD5/AMPK/HDAC4 axis.

The root of Aconitum carmichaelii Debx. (Fuzi) is an herbal medicine used in China that exerts significant efficacy in rescuing patients from severe diseases. A key toxic compound in Fuzi, aconitine (AC), could trigger unpredictable cardiotoxicities with high-individualization, thus hinders safe application of Fuzi. In this study we investigated the individual differences of AC-induced cardiotoxicities, the biomarkers and underlying mechanisms. Diversity Outbred (DO) mice were used as a genetically heterogeneous model for mimicking individualization clinically. The mice were orally administered AC (0.3, 0.6, 0.9 mg· kg-1 ·d-1) for 7 d. We found that AC-triggered cardiotoxicities in DO mice shared similar characteristics to those observed in clinic patients. Most importantly, significant individual differences were found in DO mice (variation coefficients: 34.08%-53.17%). RNA-sequencing in AC-tolerant and AC-sensitive mice revealed that hemoglobin subunit beta (HBB), a toxic-responsive protein in blood with 89% homology to human, was specifically enriched in AC-sensitive mice. Moreover, we found that HBB overexpression could significantly exacerbate AC-induced cardiotoxicity while HBB knockdown markedly attenuated cell death of cardiomyocytes. We revealed that AC could trigger hemolysis, and specifically bind to HBB in cell-free hemoglobin (cf-Hb), which could excessively promote NO scavenge and decrease cardioprotective S-nitrosylation. Meanwhile, AC bound to HBB enhanced the binding of HBB to ABHD5 and AMPK, which correspondingly decreased HDAC-NT generation and led to cardiomyocytes death. This study not only demonstrates HBB achievement a novel target of AC in blood, but provides the first clue for HBB as a novel biomarker in determining the individual differences of Fuzi-triggered cardiotoxicity.

[Exploration of transcriptome SSR markers and its application in genetic diversity assessment of Asarum sieboldii].

To explore the genetic diversity of Asarum sieboldii this study developed SSR markers based on transcriptome sequencing results and five populations of A.sieboldii from different regions were used as samples for genetic diversity assessment using software such as GenALEx 6.5, NTSYS 2.1, and Structure 2.3.4. The results showed that 16 SSR markers with high polymorphism and good repeatability were selected from the A.sieboldii transcriptome. Primers designed based on the flanking sequences of these markers successfully amplified 56 polymorphic fragments from 150 individual samples of the five A.sieboldii populations. On average, each primer amplified 3.5 polymorphic fragments, ranging from 2 to 8. The mean values of expected heterozygosity(H_e), Shannon's diversity index(I), Nei's gene diversity index(H), and the polymorphic information content(PIC) were 0.172, 0.281, 0.429, and 0.382, respectively. The mean population differentiation coefficient(F_(ST)) was 0.588, consistent with the analysis of molecular variance(AMOVA) results, which indicated greater genetic variation among A.sieboldii populations(69%) than that within populations(31%). The percentage of polymorphic loci(PPL) ranged from highest to lowest as SNJ>LN>SY>SZ>TB. Principal coordinate analysis(PCoA) and UPGMA clustering analysis further revealed genetic clustering of A.sieboldii individuals based on their geographical distribution, consistent with the results of the structure clustering analysis. In summary, the SSR markers developed from the transcriptome effectively assessed the genetic differentiation and population structure of natural A.sieboldii populations, revealing a relatively low genetic diversity in A.sieboldii, with genetic variation primarily observed at the population level and a correlation between population differentiation and geographic distance.

Pharmacological mechanisms of sinomenine in anti-inflammatory immunity and osteoprotection in rheumatoid arthritis: A systematic review.

BACKGROUND: Sinomenine (SIN) is the main pharmacologically active component of Sinomenii Caulis and protects against rheumatoid arthritis (RA). In recent years, many studies have been conducted to elucidate the pharmacological mechanisms of SIN in the treatment of RA. However, the molecular mechanism of SIN in RA has not been fully elucidated. PURPOSE: To summarize the pharmacological effects and molecular mechanisms of SIN in RA and clarify the most valuable regulatory mechanisms of SIN to provide clues and a basis for basic research and clinical applications. METHODS: We systematically searched SciFinder, Web of Science, PubMed, China National Knowledge Internet (CNKI), the Wanfang Databases, and the Chinese Scientific Journal Database (VIP). We organized our work based on the PRISMA statement and selected studies for review based on predefined selection criteria. OUTCOME: After screening, we identified 201 relevant studies, including 88 clinical trials and 113 in vivo and in vitro studies on molecular mechanisms. Among these studies, we selected key results for reporting and analysis. CONCLUSIONS: We found that most of the known pharmacological mechanisms of SIN are indirect effects on certain signaling pathways or proteins. SIN was manifested to reduce the release of inflammatory cytokines such as Tumor necrosis factor-α (TNF-α), Interleukin-6 (IL-6), and IL-1ß, thereby reducing the inflammatory response, and apparently blocking the destruction of bone and cartilage. The regulatory effects on inflammation and bone destruction make SIN a promising drug to treat RA. More notably, we believe that the modulation of α7nAChR and the regulation of methylation levels at specific GCG sites in the mPGES-1 promoter by SIN, and its mechanism of directly targeting GBP5, certainly enriches the possibilities and the underlying rationale for SIN in the treatment of inflammatory immune-related diseases.

Effect of hydrophobic property on antibacterial activities of green tea polysaccharide conjugates against Escherichia coli.

We previously found that green tea polysaccharide conjugates (gTPCs) have antibacterial activity against Escherichia coli. In this study, the effect of hydrophobic property on the antibacterial activities of gTPCs was evaluated to elucidate their property-activity relationship. Three gTPCs (gTPCs-5 h, gTPCs-12 h and gTPCs-24 h) were extracted from green tea with the ethanol precipitation time of 5 h, 12 h and 24 h, respectively. These three gTPCs did not differ significantly in terms of molecular weight distribution, amino acids composition and zeta potentials. Fourier transform infrared spectroscopy results revealed that gTPCs-5 h and gTPCs-12 h processed more hydrogen bonds than gTPCs-24 h. The surface hydrophobicity and contact angle of gTPCs-5 h were larger than that of gTPCs-12 h and gTPCs-24 h. The antibacterial activity of gTPCs against E. coli decreased in the order of gTPCs-5 h > gTPCs-12 h > gTPCs-24 h. There wasn't significant difference among the zeta potentials of E. coli treated by gTPCs-5 h, gTPCs-12 h and gTPCs-24 h, but the bacterial contact angles of E. coli treated by gTPCs-5 h were higher compared with those of the other two gTPCs. Furthermore, gTPCs-5 h exhibited higher activity to decrease bacterial membrane proteins, and increase bacterial membrane permeability than the other two gTPCs. In conclusion, gTPCs with higher hydrophobicity property exhibited stronger antibacterial activity against E. coli.

Sinomenine ameliorates collagen-induced arthritis in mice by targeting GBP5 and regulating the P2X7 receptor to suppress NLRP3-related signaling pathways.

Sinomenine (SIN) is an isoquinoline alkaloid isolated from Sinomenii Caulis, a traditional Chinese medicine used to treat rheumatoid arthritis (RA). Clinical trials have shown that SIN has comparable efficacy to methotrexate in treating patients with RA but with fewer adverse effects. In this study, we explored the anti-inflammatory effects and therapeutic targets of SIN in LPS-induced RAW264.7 cells and in collagen-induced arthritis (CIA) mice. LPS-induced RAW264.7 cells were pretreated with SIN (160, 320, 640 µM); and CIA mice were administered SIN (25, 50 and 100 mg·kg-1·d-1, i.p.) for 30 days. We first conducted a solvent-induced protein precipitation (SIP) assay in LPS-stimulated RAW264.7 cells and found positive evidence for the direct binding of SIN to guanylate-binding protein 5 (GBP5), which was supported by molecular simulation docking, proteomics, and binding affinity assays (KD = 3.486 µM). More importantly, SIN treatment markedly decreased the expression levels of proteins involved in the GBP5/P2X7R-NLRP3 pathways in both LPS-induced RAW264.7 cells and the paw tissue of CIA mice. Moreover, the levels of IL-1ß, IL-18, IL-6, and TNF-α in both the supernatant of inflammatory cells and the serum of CIA mice were significantly reduced. This study illustrates a novel anti-inflammatory mechanism of SIN; SIN suppresses the activity of NLRP3-related pathways by competitively binding GBP5 and downregulating P2X7R protein expression, which ultimately contributes to the reduction of IL-1ß and IL-18 production. The binding specificity of SIN to GBP5 and its inhibitory effect on GBP5 activity suggest that SIN has great potential as a specific GBP5 antagonist.

Caffeoyl-coenzyme A O-methyltransferase mediates regulation of carbon flux fluctuations during phenylpropenes and lignin biosynthesis in the vegetative organ roots of Asarum sieboldii Miq.

Asarum sieboldii Miq. possesses remarkable medicinal value due to its essential oil enriched with phenylpropenes (e.g., methyleugenol and safrole). Although the biosynthesis of phenylpropenes shares a common pathway with lignin, the regulation mechanisms in carbon flux allocation between them are unclear. This study is the first to genetically verify the carbon flux regulation mechanism in A. sieboldii roots. We regulated the expression of Caffeoyl-coenzyme A O-methyltransferase (CCoAOMT), an essential enzyme in the common pathway, to investigate carbon flux allocation in vegetative organs. Here, the lignin and phenylpropene content fluctuation was analyzed by wet chemistry and GC-MS methods. A bona fide CCoAOMT gene from A. sieboldii was firstly cloned and verified. Preliminary heterologous expression validation in transgenic Arabidopsis thaliana showed that RNAi-induced CCoAOMT down-regulation significantly decreased lignin content by 24% and increased the S/G ratio by 30%; however, AsCCoAOMT over-expression in A. thaliana resulted in a 40% increase in lignin content and a 20% decrease in the S/G ratio when compared to the wild type. Similar trends were noted in homologous transformation in A. sieboldii, although the variations were not conspicuous. Nevertheless, the transgenic A. sieboldii plants displayed substantial differences in the level of phenylpropene compounds methyleugenol and safrole leading to a 168% increase in the methyleugenol/safrole ratio in the over-expression line and a 73% reduction in RNAi-suppression line. These findings suggest that the biosynthesis of phenylpropene constituents methyleugenol and safrole seems to be prioritized over lignin. Furthermore, this study indicated that suppression of AsCCoAOMT resulted in marked root susceptibility to pathogenic fungal disease, implying a significant additional role of CCoAOMT in protecting plant vegetative parts from diseases. Overall, the present study provides important references and suggests that future research should be aimed at elucidating the detailed mechanisms of the carbon flux allocation between phenylpropenes and lignin biosynthesis, as well as the disease resistance competency.

Analysis of factors influencing patient delay by patients with pulmonary tuberculosis in Lishui City, Zhejiang Province.

OBJECTIVE: The purpose of this study was to collect data on the current state of patient delay by patients with tuberculosis (TB) in Lishui City, Zhejiang Province who were under the care of a TB-designated hospital from 2011 to 2021 and to analyze the factors that contribute to this problem in order to provide a scientific basis for the prevention and control of TB. METHODS: In this observational study, we collected data on patients with pulmonary TB that were reported to the Chinese government's disease prevention and control information system by the Traditional Chinese Medicine Hospital in Lishui City between 2011 and 2021. The data included demographics like age, gender, occupation, household registration, current address, date of symptoms, date of first visit, and etiology results. Multivariate logistic regression analysis was used to analyze the factors influencing patient delay by patients with pulmonary TB. RESULTS: There were 3,190 cases of pulmonary TB treated in a TB-designated hospital in Lishui City, Zhejiang Province, between 2011 and 2021. Of these, 2,268 involved patient delay, with the delay rate of 71.10% and the median (Q25, Q75) days of patient delay being 36 (25, 72) days. Results of multivariate logistic regression analysis indicated the presence of risk factors-age > 60 years old (OR = 1.367, 95% CI: 1.144 ~ 1.632), pathogen positive (OR = 1.211, 95% CI: 1.033 ~ 1.419), and employed as peasants (OR = 1.353, 95% CI:1.144 ~ 1.601) for patient delay in patients with pulmonary TB. Patients with diabetes mellitus made up 64.94% of the pulmonary TB population, which was lower than the 71.58% of patients without diabetes mellitus (χ2 = 4.602, P = 0.032). Additionally, the presence of diabetes mellitus may be a protective factor in patient delay in patients with pulmonary TB (OR = 0.641, 95% CI: 0.481 ~ 0.856). CONCLUSION: High rates of patient delay, age > 60 years old, a positive etiology, and being employed as peasants are all possible risk factors for pulmonary TB in Lishui City, Zhejiang Province.

Regulating effects of phytosterol esters-loaded emulsions stabilized with green tea polysaccharide conjugates and Tween on lipids in KKAy mice.

Phytosterol esters (PSE) have been shown to have cholesterol-lowering effects, but their insolubility in water limits their applications. Green tea polysaccharide conjugates (gTPC) have hypoglycemic and emulsifying effects. To address lipid dysregulation in diabetic patients, we developed PSE-loaded emulsions stabilized with gTPC and Tween-20 (gTPC-PSE emulsions) and evaluated their physicochemical properties. We subsequently investigated the lipid-regulating potential of these emulsions to in KKAy mice. The KKAy mice were randomly assigned to eight groups: the model group, the Lipitor (10 mg·kg-1)-acarbose (30 mg·kg-1) combination group, two gTPC groups, two PSE groups, and two gTPC-PSE groups with a 1:2 mass ratio of gTPC to PSE. The administered doses were 90 and 270 mg kg-1, respectively. Administration of a 270 mg·kg-1 dose of gTPC-PSE emulsions led to the most significant effects including increased levels of liver and serum high-density lipoprotein cholesterol (HDL-CH), reduced serum leptin and insulin, and improved liver superoxide dismutase (SOD) and reduced malondialdehyde (MDA). In general, gTPC and PSE demonstrated a synergistic effect on lipid regulation in mice. Our results indicate that gTPC-PSE emulsions hold potential as a nutritional intervention for diabetes by modulating lipid levels.

Assessment of Effective Anti-TB Regimens and Adverse Outcomes Related Risk Factors in the Elderly and Senile-Aged TB Patients.

Objective: Compared to younger patients with tuberculosis (TB), elderly and senile-aged patients with TB had a higher incidence of adverse outcomes particularly in terms of lost to follow-up and deaths. Our study aimed to gain insight into the effectiveness of anti-tuberculosis (anti-TB) treatment in the elderly or senile-aged patients and identify the risk factors for adverse outcomes. Methods: The case information was obtained from the "Tuberculosis Management Information System". From January 2011 to December 2021, this retrospective analysis was conducted in Lishui City, Zhejiang Province to observe and record the outcomes of elderly patients diagnosed with TB who agreed to receive anti-TB and(or) traditional Chinese medicine(TCM) treatment. We also employed a logistic regression model to analyze the risk factors for adverse outcomes. Results: Among the 1191 elderly or senile-aged patients with TB who received the treatment, the success rate was 84.80% (1010/1191). Using logistic regression analysis, several risk factors for adverse outcomes (failure, death, loss to follow-up) were identified, including age ≥ 80 years (OR 2.186, 95% CI 1.517~3.152, P<0.001), lesion area ≥ 3 lung fields (OR 0.410, 95% CI 0.260~0.648, P<0.001), radiographic lesions failing to improve after 2 months of treatment (OR 2.048, 95% CI 1.302~3.223, P=0.002), sputum bacteriology failing to turn negative after 2 months of treatment (OR 2.213, 95% CI 1.227~3.990, P=0.008), lack of a standardized treatment plan (OR 2.095, 95% CI 1.398~3.139, P<0.001), and non-involvement of traditional Chinese medicine (OR 2.589, 95% CI 1.589~4.216, P<0.001). Conclusion: The anti-TB treatment success rate in the elderly and senile-aged patients is suboptimal. Contributing factors include advanced age, extensive lesions, and low sputum negative conversion rate during the intensive treatment phase. The results will informative and could be useful for policy maker for to control of reemergence of TB in big cities.

Genomic Resequencing Unravels the Genetic Basis of Domestication, Expansion, and Trait Improvement in Morus Atropurpurea.

Mulberry is an economically important plant in the sericulture industry and traditional medicine. However, the genetic and evolutionary history of mulberry remains largely unknown. Here, this work presents the chromosome-level genome assembly of Morus atropurpurea (M. atropurpurea), originating from south China. Population genomic analysis using 425 mulberry accessions reveal that cultivated mulberry is classified into two species, M. atropurpurea and M. alba, which may have originated from two different mulberry progenitors and have independent and parallel domestication in north and south China, respectively. Extensive gene flow is revealed between different mulberry populations, contributing to genetic diversity in modern hybrid cultivars. This work also identifies the genetic architecture of the flowering time and leaf size. In addition, the genomic structure and evolution of sex-determining regions are identified. This study significantly advances the understanding of the genetic basis and domestication history of mulberry in the north and south, and provides valuable molecular markers of desirable traits for mulberry breeding.

A comprehensive review of polyphenol oxidase in tea (Camellia sinensis): Physiological characteristics, oxidation manufacturing, and biosynthesis of functional constituents.

Polyphenol oxidase (PPO) is a metalloenzyme with a type III copper core that is abundant in nature. As one of the most essential enzymes in the tea plant (Camellia sinensis), the further regulation of PPO is critical for enhancing defensive responses, cultivating high-quality germplasm resources of tea plants, and producing tea products that are both functional and sensory qualities. Due to their physiological and pharmacological values, the constituents from the oxidative polymerization of PPO in tea manufacturing may serve as functional foods to prevent and treat chronic non-communicable diseases. However, current knowledge of the utilization of PPO in the tea industry is only available from scattered sources, and a more comprehensive study is required to reveal the relationship between PPO and tea obviously. A more comprehensive review of the role of PPO in tea was reported for the first time, as its classification, catalytic mechanism, and utilization in modulating tea flavors, compositions, and nutrition, along with the relationships between PPO-mediated enzymatic reactions and the formation of functional constituents in tea, and the techniques for the modification and application of PPO based on modern enzymology and synthetic biology are summarized and suggested in this article.

Oolong tea of different years protects high-fat diet-fed mice against obesity by regulating lipid metabolism and modulating the gut microbiota.

Long-term stored oolong tea has recently attracted considerable attention concerning its salutary effect. In this study, the anti-obesity effect of different years' oolong tea on high-fat diet-fed mice was compared. Wuyi rock tea of 2001, 2011, and 2020 were chosen to be the representative samples of oolong tea. The results showed that eight-week administration of 2001 Wuyi rock tea (WRT01), 2011 Wuyi rock tea (WRT11), and 2020 Wuyi rock tea (WRT20) extracts (400 mg per kg per d) significantly decreased the body weight and attenuated the obesity in high-fat diet-fed mice. 2001 and 2011 Wuyi rock teas reduced obesity mainly through regulating lipid metabolism and activating the AMPK/SREBP-1 pathway, downregulating the expression of SREBP-1, FAS, and ACC and upregulating CPT-1a expression; while the 2011 and 2020 Wuyi rock teas by moderating the gut microbiota dysbiosis, reshaping the gut microbiota, and promoting the growth of beneficial bacteria, especially Akkermansia. 2011 Wuyi rock tea was proven to be more effective in reducing body weight gain and liver oxidative stress than the others. Collectively, all three Wuyi rock teas of different years alleviated high-fat diet-induced obesity by regulating lipid metabolism and modulating gut microbiota, whereas the emphasis of their internal mechanism is different with different storage ages.

Shouhui Tongbian Capsules induce regression of inflammation to improve intestinal barrier in mice with constipation by targeted binding to Prkaa1: With no obvious toxicity.

Constipation arising from the poor bowel movement is a rife enteric health problem. Shouhui Tongbian Capsule (SHTB) is a traditional Chinese medicine (TCM) which effectively improve the symptoms of constipation. However, the mechanism has not been fully evaluated. The purpose of this study was to evaluate the effect of SHTB on the symptoms and intestinal barrier of mice with constipation. Our data showed that SHTB effectively improved the constipation induced by diphenoxylate, which was confirmed by shorter first defecation time, higher internal propulsion rate and fecal water content. Additionally, SHTB improved the intestinal barrier function, which was manifested by inhibiting the leakage of Evans blue in intestinal tissues and increasing the expression of occludin and ZO-1. SHTB inhibited NLRP3 inflammasome signaling pathway and TLR4/NF-κB signaling pathway, reduced the number of proinflammatory cell subsets and increased the number of immunosuppressive cell subsets to relieve inflammation. The photochemically induced reaction coupling system combined with cellular thermal shift assay and central carbon metabolomics technology confirmed that SHTB activated AMPKα through targeted binding to Prkaa1 to regulate Glycolysis/Gluconeogenesis and Pentose Phosphate Pathway, and finally inhibited intestinal inflammation. Finally, no obvious toxicity related to SHTB was found in a repeated drug administration toxicity test for consecutive 13 weeks. Collectively, we reported SHTB as a TCM targeting Prkaa1 for anti-inflammation to improve intestinal barrier in mice with constipation. These findings broaden our knowledge of Prkaa1 as a druggable target protein for inflammation inhibition, and open a new avenue to novel therapy strategy for constipation injury.

Salvianolic acid B suppresses hepatic fibrosis by inhibiting ceramide glucosyltransferase in hepatic stellate cells.

UDP-glucose ceramide glucosyltransferase (UGCG) is the first key enzyme in glycosphingolipid (GSL) metabolism that produces glucosylceramide (GlcCer). Increased UGCG synthesis is associated with cell proliferation, invasion and multidrug resistance in human cancers. In this study we investigated the role of UGCG in the pathogenesis of hepatic fibrosis. We first found that UGCG was over-expressed in fibrotic livers and activated hepatic stellate cells (HSCs). In human HSC-LX2 cells, inhibition of UGCG with PDMP or knockdown of UGCG suppressed the expression of the biomarkers of HSC activation (α-SMA and collagen I). Furthermore, pretreatment with PDMP (40 µM) impaired lysosomal homeostasis and blocked the process of autophagy, leading to activation of retinoic acid signaling pathway and accumulation of lipid droplets. After exploring the structure and key catalytic residues of UGCG in the activation of HSCs, we conducted virtual screening, molecular interaction and molecular docking experiments, and demonstrated salvianolic acid B (SAB) from the traditional Chinese medicine Salvia miltiorrhiza as an UGCG inhibitor with an IC50 value of 159 µM. In CCl4-induced mouse liver fibrosis, intraperitoneal administration of SAB (30 mg · kg-1 · d-1, for 4 weeks) significantly alleviated hepatic fibrogenesis by inhibiting the activation of HSCs and collagen deposition. In addition, SAB displayed better anti-inflammatory effects in CCl4-induced liver fibrosis. These results suggest that UGCG may represent a therapeutic target for liver fibrosis; SAB could act as an inhibitor of UGCG, which is expected to be a candidate drug for the treatment of liver fibrosis.

MOF-implanted poly (acrylamide-co-acrylic acid)/chitosan organic hydrogel for uranium extraction from seawater.

In this study, a composite hydrogel with a low swelling ratio, excellent mechanical properties, and good U (VI) adsorption capacity was developed by incorporating a metal-organic framework (MOF) with a poly (acrylamide-co-acrylic acid)/chitosan (P(AM-co-AA)/CS) composite. The CS chain, which contains NH2, reduces the swelling ratio of the hydrogel to 4.17 after 5 h of immersion in water. The coordinate bond between the MOF and carboxyl group on the surface of P(AM-co-AA)/CS improves the mechanical properties and stability of P(AM-co-AA)/CS. The U(VI) adsorption capacity of P(AM-co-AA)/CS/MOF-808 is 159.56 mg g-1 at C0 = 99.47 mg L-1 and pH = 8.0. The adsorption process is well fitted by the Langmuir isotherm and pseudo-second-order model. The P(AM-co-AA)/CS/MOF-808 also exhibits good repeatability and stability after five adsorption-desorption cycles. The uranium adsorption capacity of the developed adsorbent after one month in natural seawater is 6.2 mg g-1, and the rate of uranium adsorption on the hydrogel is 0.21 mg g-1 day-1.

Shen-Qi-Jiang-Tang granule ameliorates diabetic nephropathy via modulating tumor necrosis factor signaling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Shen-Qi-Jiang-Tang granule (SQJTG), a classic traditional Chinese medicine (TCM) prescription, has been widely used in clinical for diabetes, especially type Ⅱ diabetes. Previous anti-diabetic studies stumbled across that SQJTG has a potential kidney protective effect on diabetic nephropathy (DN). However, the protective mechanism of SQJTG on DN still needs to be explored. AIM OF THE STUDY: The purpose of the present study was to explore the therapeutic effect of SQJTG on DN through both bioinformatics analysis and in vivo experiments. METHODS AND MATERIALS: The TCMIP database was used for screening potential compounds and targets of SQJTG, and the GeneCards, OMIM, DrugBank, and TTD databases were used for collecting DN-related genes. Then protein-protein interaction analysis for the common targets of SQJTG and DN was performed by the STRING database. Meanwhile, KEGG and GO were carried out using the Metascape and DAVID databases. In vivo experiments, to testify the potential kidney protective effects of SQJTG, STZ-induced DN mice with different dosages of SQJTG treatment were collected and the renal tissues were detected by H&E, PAS, Masson and TUNEL staining. Immunohistochemistry and immunoblotting were used to assess the proteins' expressions. Flow cytometry and ELISA assay were used to detect the levels of pro-inflammatory cytokines. RESULTS: Among the 338 compounds ascertained by SQJTG, there were 789 related targets as well. Moreover, 1,221 DN-related targets were predicted and 20 core targets were screened by the PPI analyses. According to GO and KEGG pathway analysis, SQJTG may affect DN via the TNF pathway. For the in vivo experiments, renal histomorphological examinations demonstrated that SQJTG treatment significantly ameliorated STZ-induced kidney damage and had a dosage dependence. Meanwhile, mice with DN were found to have dramatic increases in IL-1, TNF-α, IL-6, and IL-12, but markedly decreased after administration of SQJTG. In addition, the protein levels of TNF signaling molecules, like p-P65, p-JNK, and p-p38, showed significantly elevated in kidney tissues of DN mice and attenuated after SQJTG treatment. CONCLUSIONS: SQJTG exerts a kidney protective effect in DN mice via modulating TNF signaling pathways, and it has promising applications for the treatment of DN.

Molecular cloning and characterization of Cinnamoyl-CoA reductase promoter gene from Asarum sieboldii Miq.

Asarum sieboldii Miq., a perennial herb of the family Aristolochiaceae, is widely used in China to treat cold, fever, aphthous stomatitis, toothache, gingivitis, and rheumatoid arthritis. Methyleugenol is the most representative pharmacological constituent of this medicinal herb. Cinnamoyl-CoA reductase (CCR), which has been well known for occupying a critical position in the lignin biosynthesis pathway, is also shared with the biosynthesis of methyleugenol. To better understand the regulatory mechanisms of methyleugenol biosynthesis, a 1530-bp long promoter region of the AsCCR1 gene was isolated. PLACE and PlantCARE analysis affirmed the existence of the core promoter elements such as TATA and CAAT boxes, abiotic stress-responsive cis-regulation elements like abscisic acid-responsive element, G-box, and MBS in the isolated sequence. The histochemical assay suggested that it was a constitutive promoter, highly expressed in the root tissue. Moreover, the region of -200 bp to ATG (start codon) was enough to drive the expression of It GUS gene. Treatments with low temperature and high concentration of gibberellin or abscisic acid demonstrated the abiotic stress-induced expression of the AsCCR1 promoter. Overall, this study revealed the isolation and characterization of the AsCCR1 promoter. Moreover, it also provided a candidate gene for molecular breeding in A. sieboldii to enhance its pharmacological potential.

Characterization of Angraecum (Angraecinae, Orchidaceae) Plastomes and Utility of Sequence Variability Hotspots.

Angraecum, commonly known as Darwin's orchid, is the largest genus of Angraecinae (Orchidaceae). This genus exhibits a high morphological diversity, making it as a good candidate for macroevolutionary studies. In this study, four complete plastomes of Angraecum were firstly reported and the potential variability hotspots were explored. The plastomes possessed the typical quadripartite structure and ranged from 150,743 to 151,818 base pair (bp), with a guanine-cytosine (GC) content of 36.6-36.9%. The plastomes all contained 120 genes, consisting of 74 protein-coding genes (CDS), 38 transfer RNA (tRNA) genes and 8 ribosomal RNA (rRNA) genes; all ndh genes were pseudogenized or lost. A total of 30 to 46 long repeats and 55 to 63 SSRs were identified. Relative synonymous codon usage (RSCU) analysis indicated a high degree of conservation in codon usage bias. The Ka/Ks ratios of most genes were lower than 1, indicating that they have undergone purifying selection. Based on the ranking of Pi (nucleotide diversity) values, five regions (trnSGCU-trnGGCC, ycf1-trnNGGU, trnNGUU-rpl32, psaC-ndhE and trnSGCU-trnGGCC) and five protein-coding genes (rpl32, rps16, psbK, rps8, and ycf1) were identified. The consistent and robust phylogenetic relationships of Angraecum were established based on a total of 40 plastomes from the Epidendroideae subfamily. The genus Angraecum was strongly supported as a monophyletic group and sister to Aeridinae. Our study provides an ideal system for investigating molecular identification, plastome evolution and DNA barcoding for Angraecum.

UCP2-dependent redox sensing in POMC neurons regulates feeding.

Paradoxically, glucose, the primary driver of satiety, activates a small population of anorexigenic pro-opiomelanocortin (POMC) neurons. Here, we show that lactate levels in the circulation and in the cerebrospinal fluid are elevated in the fed state and the addition of lactate to glucose activates the majority of POMC neurons while increasing cytosolic NADH generation, mitochondrial respiration, and extracellular pyruvate levels. Inhibition of lactate dehydrogenases diminishes mitochondrial respiration, NADH production, and POMC neuronal activity. However, inhibition of the mitochondrial pyruvate carrier has no effect. POMC-specific downregulation of Ucp2 (Ucp2PomcKO), a molecule regulated by fatty acid metabolism and shown to play a role as transporter in the malate-aspartate shuttle, abolishes lactate- and glucose-sensing of POMC neurons. Ucp2PomcKO mice have impaired glucose metabolism and are prone to obesity on a high-fat diet. Altogether, our data show that lactate through redox signaling and blocking mitochondrial glucose utilization activates POMC neurons to regulate feeding and glucose metabolism.

TET3 epigenetically controls feeding and stress response behaviors via AGRP neurons.

The TET family of dioxygenases promote DNA demethylation by oxidizing 5-methylcytosine to 5-hydroxymethylcytosine (5hmC). Hypothalamic agouti-related peptide-expressing (AGRP-expressing) neurons play an essential role in driving feeding, while also modulating nonfeeding behaviors. Besides AGRP, these neurons produce neuropeptide Y (NPY) and the neurotransmitter GABA, which act in concert to stimulate food intake and decrease energy expenditure. Notably, AGRP, NPY, and GABA can also elicit anxiolytic effects. Here, we report that in adult mouse AGRP neurons, CRISPR-mediated genetic ablation of Tet3, not previously known to be involved in central control of appetite and metabolism, induced hyperphagia, obesity, and diabetes, in addition to a reduction of stress-like behaviors. TET3 deficiency activated AGRP neurons, simultaneously upregulated the expression of Agrp, Npy, and the vesicular GABA transporter Slc32a1, and impeded leptin signaling. In particular, we uncovered a dynamic association of TET3 with the Agrp promoter in response to leptin signaling, which induced 5hmC modification that was associated with a chromatin-modifying complex leading to transcription inhibition, and this regulation occurred in both the mouse models and human cells. Our results unmasked TET3 as a critical central regulator of appetite and energy metabolism and revealed its unexpected dual role in the control of feeding and other complex behaviors through AGRP neurons.