Multiomics reveals the ameliorating effect and underlying mechanism of aqueous extracts of polygonatum sibiricum rhizome on obesity and liver fat accumulation in high-fat diet-fed mice.

BACKGROUND: Polygonatum sibiricum polysaccharides protect against obesity and NAFLD. However, the potential effects of PS rhizome aqueous extracts (PSRwe) on adiposity and hepatic lipid accumulation remains unexplored. PURPOSE: Elucidating the impact and underlying mechanism of PSRwe on HFD-induced obesity and liver fat depostition. STUDY DESIGN: 56 male mice, aged eight weeks, were divided into seven groups: Positive, four doses of PSRwe, Model, and Control. HFD was fed for eight weeks, followed by alternate-day gavage of orlistat and PSRwe for an additional eight-week period. Integrative analysis encompassing multiomics, physiological and histopathological, and biochemical indexes was employed. METHODS: Body weight (BW); liver, fat and Lee's indexes; TC, TG, LDL-C, HDL-C, AST, ALT, FFA, leptin, and adiponectin in the liver and blood; TNFα, IL-6, and LPS in the colon, plasma, and liver; H&E, PAS and oil red O staining on adipose and liver samples were examined. OGTT and ITT were conducted The gut microbiome, microbial metabolome, colonic and liver transcriptome, plasma and liver metabolites were investigated. RESULTS: PSRwe at the dosage of 7.5 mg/kg demonstrated significant and consistent reduction in BW and hepatic fat deposition than orlistat. PSRwe significantly decreased TC, TG, LDL-C, LEP, FFA levels in blood and liver. PSRwe significantly enhanced the relative abundance of probiotics including Akkermansia muciniphila, Bifidobacterium pseudolongum, Lactobacillus reuteri, and metabolic pathways including glycolysis and fatty acids ß-oxidation. The 70 up-regulated microbial metabolites in PSRwe-treated mice mainly involved in nucleotides and amino acids metabolism, while 40 decreased metabolites primarily associated with lipid metabolism. The up-regulated colonic differentially expressed genes (DEGs) participate in JAK-STAT/PI3K-Akt/FoxO signaling pathway, serotonergic/cholinergic/glutamatergic synapses, while the down-regulated DEGs predominantly focused on fat absorption and transport. The up-regulated liver DEGs mainly concentrated on fatty acid oxidation and metabolism. Liver metabolisms revealed 131 differential metabolites, among which carnitine and oxidized lipids significantly increased in PSRwe-treated mice. In plasma, the 58 up-regulated metabolites mainly participate in co-factors/vitamins metabolism while 154 down-regulated ones in fatty acids biosynthesis. Comprehensive multiomics association analysis revealed significant associations between gut microbiota and colonic/liver gene expression, and suggested exogenous and endogenous betaine may be active compound in alleviating HFD-induced symptoms. CONCLUSION: PSRwe effectively mitigate HFD-induced obesity and hepatic steatosis by increasing beneficial bacteria, reducing colonic fat digestion/absorption, increasing hepatic lipid metabolism, and elevating betaine levels.

Effect of ultrasonic degradation on the physicochemical characteristics, GLP-1 secretion, and antioxidant capacity of Polygonatum cyrtonema polysaccharide.

This study aimed to evaluate the influence of ultrasonic degradation on the physicochemical and biological characteristics of Polygonatum cyrtonema polysaccharide (PCP, 8.59 kDa). PCP was subjected to ultrasonic treatment for 8, 16, and 24 h and yielded the degraded fractions PCP-8, PCP-16, and PCP-24 (5.06, 4.13, and 3.69 kDa), respectively. Compared with the intact PCP, PCP-8, PCP-16 and PCP-24 had a reduced particle size (decrements of 28.03 %, 46.15 % and 62.54 %, respectively). Although ultrasonic degradation did not alter the primary structure of PCP, its triple helical and superficial structures were disrupted, with degraded fractions demonstrating reduced thermal stability and apparent viscosities compared with those of the intact PCP. Furthermore, the functional properties of the degraded fractions were different. PCP-16 most favourably affected GLP-1 secretion, while PCP-8 and PCP-24 exhibited the strongest antioxidant and enzyme inhibitory activities, respectively. Hence, controlled ultrasound irradiation is an appealing approach for partially degrading PCP and enhancing its bioactivity as a functional agent.

Polygonatum cyrtonema polysaccharides reshape the gut microbiota to ameliorate dextran sodium sulfate-induced ulcerative colitis in mice.

Ulcerative colitis (UC) is a chronic inflammatory bowel disease characterized inflammatory imbalance, intestinal epithelial mucosal damage, and dysbiosis of the gut microbiota. Polygonatum cyrtonema polysaccharides (PCPs) can regulate gut microbiota and inflammation. Here, the different doses of PCPs were administered to dextran sodium sulfate-induced UC mice, and the effects of the whole PCPs were compared with those of the fractionated fractions PCP-1 (19.9 kDa) and PCP-2 (71.6 and 4.2 kDa). Additionally, an antibiotic cocktail was administered to UC mice to deplete the gut microbiota, and PCPs were subsequently administered to elucidate the potential role of the gut microbiota in these mice. The results revealed that PCP treatment significantly optimized the lost weight and shortened colon, restored the balance of inflammation, mitigated oxidative stress, and restored intestinal epithelial mucosal damage. And, the PCPs exhibited superior efficacy in ameliorating these symptoms compared with PCP-1 and PCP-2. However, depletion of the gut microbiota diminished the therapeutic effects of PCPs in UC mice. Furthermore, fecal transplantation from PCP-treated UC mice to new UC-afflicted mice produced therapeutic effects similar to PCP treatment. So, PCPs significantly ameliorated the symptoms, inflammation, oxidative stress, and intestinal mucosal damage in UC mice, and gut microbiota partially mediated these effects.

Transcriptomic Analysis Reveals the Flavonoid Biosynthesis Pathway Involved in Rhizome Development in Polygonatum cyrtonema Hua.

Polygonatum cyrtonema Hua (P. cyrtonema) rhizomes are rich in flavonoids and other secondary metabolites, exhibiting remarkable antioxidant, anti-tumor, and immunomodulatory effects. Polygonatum flavonoid-biosynthesis-related genes have been characterized already. However, a comprehensive overview of Polygonatum flavonoid biosynthesis pathways is still absent. To articulate the accumulation of the flavonoid biosynthesis pathways, we examined transcriptome changes using Illumina HiSeq from five different tissues and the RNA-seq of 15 samples had over 105 Gb of a clean base, generating a total of 277,955 unigenes. The cDNA libraries of the fruits (F), leaves (L), roots (R), stems (S), and rhizomes (T) of three-year-old P. cyrtonema plants generated 57,591, 53,578, 60,321, 51,530, and 54,935 unigenes. Comparative transcriptome analysis revealed that 379 differentially expressed genes (DEGs) were in the group of F _vs_ T, L _vs_ T, R _vs_ T, and S _vs_ T, and the transcripts of flavonoid-biosynthesis-related DEGs were principally enriched in rhizomes. In addition, combined with WGCNA and the FPKM of five tissues' transcription, nine differentially expressed transcription factor families (MYB, WRKY, AP2/ERF, etc.) were characterized in the red module, the red module positively correlated with rhizome flavonoid accumulation. Quantitative real-time PCR (qRT-PCR) further indicated that BZIP1, C3H31, ERF114, and DREB21 are differentially expressed in rhizomes, accompanied in rhizome development in P. cyrtonema. Therefore, this study provides a foundation for further research into uncovering the accumulation of flavonoid biosynthesis in the rhizomes of P. cyrtonema.

The complete chloroplast genomes of Polygonatum hunanense, P. verticillatum, and P. caulialatum and their phylogenetic positions.

Polygonatum hunanense H.H. Liu & B.Z. Wang (2021) and P. verticillatum (L.) All. (1875) have been widely used as foods and as folk medicines in China and India, and P. caulialatum S. R. Yi (2021) has recently been described as a new medical plant in China. There is at present a lack of genome information regarding the species. Hence, this study reports the complete chloroplast genomes of the three species. The genomes of P. hunanense, P. verticillatum, and P. caulialatum were 155,583 bp, 155,650 bp, and 155,352 bp in length, respectively. They contained large single-copy (LSC) regions of 84,412 bp, 84,404 bp, and 84,285 bp, small single-copy (SSC) regions of 18,427 bp, 18,416 bp, and 18,463 bp, and a pair of inverted repeats of 26,372 bp, 26,415 bp, and 26,302 bp, respectively. The chloroplast genomes of P. hunanense, P. verticillatum, and P. caulialatum had 133 (103 unique) genes, consisting of 87 protein-coding genes, 38 ribosomal ribonucleic acid (RNA) genes, and eight transfer RNA genes, respectively. A maximum-likelihood phylogenetic tree showed that P. kingianum Coll. et Hemsl. var. grandifolium D.M. Liu & W.Z. Zeng (1991) was closer to P. cyrtonema Hua (1892) rather than to P. kingianum Coll. et Hemsl. (1890), further supporting its status as a unique species of the genus. Moreover, P. verticillatum was separated from the easily confused herb P. cirrhifolium (Wall.) Royle (1839), while P. caulialatum was closest to P. humile Fisch. ex Maxim. (1859). This research provides a foundation for further study of these herbs.

Quality analysis and function prediction of soil microbial communities of Polygonatum cyrtonema in two indigenous-origins.

Polygonatum cyrtonema Hua (PCH), as an important economic crop, is used as raw industrial materials and traditional Chinese medicine. There are significant variations in the quality of PCH from different geographical origins. It can be due to the change of the endophytic fungi and soil microbial communities of PCH. Therefore, the aim of this study is to investigate the composition and functional prediction of the main microbial communities in the rhizomes and soil of PCH and explore their impact on medicinal quality. High-throughput sequencing techniques targeting ITS and 16S rDNA were employed to compare the structure and biodiversity differences of endophytic fungi in the rhizomes and soil microbial communities of PCH from 12 different locations in Sichuan and Guangxi province. Heatmap analysis was used for comprehensive statistics and visualization of the richness of rhizome and soil microbial communities from all locations. Venn analysis was conducted to determine the total number of shared fungi between rhizomes and soil, and GraphPad Prism analysis was employed to predict and compare the microbial communities related to phenotypes at the genus level in Sichuan and Guangxi. Tax4Fun and Fungild were used for metabolic function prediction of microbial communities in the rhizomes and soil of PCH. The results revealed the identification of 19,387 bacterial amplicon sequence variants (ASVs) in the rhizomes and 37,990 bacterial ASVs in the soil, with 6,889 shared bacterial ASVs. In addition, 2,948 fungal ASVs were identified in the rhizomes and 8,868 in the soil, with 1,893 shared fungal ASVs. Microbial sequencing results indicated that the fungal communities between soil and rhizomes were mainly composed of Ascomycota and Basidiomycota, while bacterial communities included Proteobacteria, Acidobacteria, Bacteroidota, Gammatimonadota, and Firmicutes. Dominant bacterial groups such as Nitrospira, Acidibacter, and fungal groups including Mortierella, Ceratobasidium, and Fusarium were identified as potential contributors to the observed traits. In the top 15 microbial genera, both Sichuan and Guangxi contain 15 bacterial genera, but there are differences in their abundance. Guangxi has three unique fungal genera, including the genera Scleroderma, Russula, and Gliocladiopsis. On the other hand, Sichuan has the unique fungal genus Chamaeota. The correlation analysis between the microbiota and the chemical content from 12 different collecting spots was performed by GraphPad Prism. Burkholderia-Caballeronia-Paraburkholderia, Acidibacter, and Amycolatopsis show an inverse proportionality to total polysaccharides and saponins, while Enterobacter shows a direct proportionality to total polysaccharides and inverse proportionality to saponins. The metabolism pathways show a significant positive correlation with PCH polysaccharides and saponins. This study provide new insights into the mechanisms underlying the quality differences between the two major indigenous areas.

Polygonatum polysaccharide ameliorates D-galactose-induced cognitive dysfunction in aging rats by inhibiting ferroptosis through activation of Nrf2.

CONTEXT: Aging is a major risk factor for various neurodegenerative diseases, and ferroptosis has been identified as an important mode of cell death during accelerated aging. As the main component of the edible plant YuZhu in China, Polygonatum polysaccharide (POP) is an important natural compound with anti-aging properties. OBJECTIVE: To evaluate the anti-aging effects of POP and the underlying molecular mechanisms involved and to evaluate the overall anti-aging effects of POP on cognitive impairment due to accelerated aging. MATERIALS AND METHODS: A D-galactose (D-gal)-induced accelerated aging rat model was established to evaluate the anti-aging effects of POP and the underlying molecular mechanisms involved. In turn, Morris water maze and open field experiments were used to evaluate the anti-aging effects of POP on cognitive impairment due to accelerated aging. RESULTS: The mechanism by which POP affects nuclear factor E2-related factor 2 (Nrf2), an essential transcription factor, was confirmed. POP significantly improved d-gal-induced cognitive dysfunction in treated model rats, which exhibited reduced pathological changes in the hippocampus, reduced latency of the water maze platform, and increased exploration time in the central area in the open field experiment compared to those of untreated model rats. Furthermore, POP intervention downregulated ferroptosis-related proteins and upregulated Nrf2 expression, and selective inhibition of Nrf2 eliminated the ability of POP to reduce ferroptosis. CONCLUSIONS: POP is a natural ingredient with therapeutic potential due to its ability to alleviate aging by activating Nrf2, inhibiting ferroptosis, and alleviating cognitive dysfunction.

First Report of Root Rot on Rhizome of Polygonatum kingianum Caused by Aspergillus awamori.

Polygonatum kingianum Coll. et Hemsl (Huangjing), which belongs to the family Asparagaceae, is a perennial traditional Chinese herb with homologous medicinal and edible value (Liu et al., 2021). Huangjing is known to promote blood circulation; it has anti-inflammatory properties, increases immunity, and provides hypoglycemic treatments (Ma et al., 2019). Root rot-infected P. kingianum exhibited withering yellow leaves and stems, rhizome rot, slowed growth, and plant death. In recent years, with an average incidence of up to 45%, the spread of HJ root rot (rhizome and stem bases) has resulted in a significant reduction in the quality and up to 63% reduction in the yields of Sichuan Junlian (104.5°E, 28.2°N) and Guizhou Zhunyi (107.0°E, 27.7°N). After collecting the diseased samples, we used the tissue isolation method to isolate the pathogenic fungi (Wu et al., 2020). Four fungal isolates associated with root rot were obtained: HJ-G2 (two strains), HJ-G3 (one strain), HJ-G4 (one strain), and HJ-G6 (two strains), of which HJ-G2 and HJ-G6 were the dominant species. To determine pathogenicity of each strain, tests were conducted by wounding rhizomes wth an inoculation needle and the pathogen strain was inoculated onto the wound and symptoms observed. The results reveal that HJ-G6 exhibited the strongest pathogenicity against P. kingianum (Figure 1). The HJ-G6 colonies were black, grew rapidly, and produced a large number of spores (Figure 1). A spherical apical sac (conidial head) is formed at the top with two palisades of cells, metulae and phialides, which are shaped radially and produce a large number of spores with 2-5 um in diameter (Figure 2). Morphological observations revealed that the isolate was consistent with Aspergillus awamori (Naher et al., 2021). To further confirm the fungal species, the ribosomal internal transcribed spacer (ITS), ß-tubulin (TUB), and elongation factor 1-alpha (EF-1a) gene regions were amplified with ITS1/ITS4, Bt2a/ Bt2b, and EF1/EF2. Primer and PCR amplification were performed as previously described (Paul et al., 2017). The sequences were compared with those obtained from GenBank. The ITS sequences (GenBank accession number OR682143) of the isolates (HJ-G6) were 100% identical to those of the strain PANCOM10 (GenBank accession number MT007535.1) of Aspergillus awamori. The EF-1a sequences (GenBank accession OR752352) of the isolates (HJ-G6) were 98% identical with strain ITEM 4777 (GenBank accession FN665402.1) of Aspergillus awamori. The TUB sequences (GenBank accession number OR752351) of the isolate (HJ-G6) were 100% identical with strain AF158 (GenBank accession MH781275.1) of Aspergillus awamori. Three maximum likelihood trees were constructed using MEGA v5.0 (Kumar et al., 2018) based on the sequences (ITS, TUB, and EF-1a) of the HJ-G6 strain and that of Aspergillus spp. previously deposited in GenBank (Paul et al., 2017). Phylogenetic analysis showed that HJ-G6 belonged to the Aspergillus awamori clade (Figure 3). Combined with morphological analysis and DNA sequencing, HJ-G6 was identified as Aspergillus awamori. To verify pathogenicity, P. kingianum roots were inoculated with the colonized agar discs of the isolates. P. kingianum plants inoculated with uncolonized agar discs were used as controls. After inoculation, P. kingianum roots were moved to the inoculation chamber under high humidity at 28 °C for 1 d and then transferred to a greenhouse. Previous studies have reported that Fusarium sp. are root rot pathogens in the rhizomes of medicinal plants (Pang et al., 2022; Song et al., 2023). In this study, HJ-G2, HJ-G3, and HJ-G4 were used as the positive controls. Typical symptoms of root rot appeared 3 days after inoculation and were similar to those observed in the field, whereas the control plants remained symptomless. According to the results of the inoculation experiment, the pathogenicity of Aspergillus awamori to P. kingianum root rot was significantly stronger than that of Fusarium (Figure 1). The pathogen was isolated from the rotting root of P. kingianum and the ITS region was sequenced again. Alignment analysis of the ITS sequences revealed that the causal agents were consistent with those of the original isolates. These studies fulfill Koch's postulates. As far as we know, this is the first report of Aspergillus awamori causing root rot in P. kingianum.

Polygonatum sibiricum polysaccharides protect against knee osteoarthritis by inhibiting the TLR2/NF-κB signaling pathway in vivo and in vitro.

Polygonatum sibiricum polysaccharides (PSP), the primary constituent of Polygonatum sibiricum, have been shown to exhibit a wide range of pharmacological effects, but their impact on osteoarthritis (OA) remains unclear. The objective of this study was to investigate the protective effects of PSP against OA and to elucidate its underlying molecular mechanism. In our in vitro experiments, PSP not only inhibited the IL-1ß-induced inflammatory responses and the nuclear factor kappa-B (NF-κB) signaling pathway in chondrocytes but also regulated the cartilage matrix metabolism. In addition, we detected 394 significantly differentially expressed genes through RNA-seq analysis on PSP-intervened chondrocytes, and the toll-like receptor 2 (TLR2) was identified as the most important feature by functional network analysis and qRT-PCR. It was also revealed that PSP treatment significantly reversed the IL-1-induced up-regulation of TLR2 expression in chondrocytes, while TLR2 overexpression partially inhibited the regulatory effects of PSP on inflammation, NF-κB signaling pathway and matrix metabolism. In our in vivo experiments, PSP treatment alleviated the development of destabilization of medial meniscus (DMM)-induced OA in mouse knee joints, inhibited the DMM-induced activation of the TLR2/NF-κB signaling pathway in mouse knee joint cartilage, and reduced the serum levels of inflammatory cytokines. In conclusion, PSP exerts its anti-inflammatory, matrix synthesis-promoting and matrix catabolism-suppressing effects in knee OA by inhibiting the TLR2/NF-κB signaling pathway, suggesting that PSP may be potentially targeted as a novel all-natural, low-toxicity drug for OA prevention and treatment.

Identification of the inhibition mechanism of carbonic anhydrase II by fructooligosaccharides.

Polygonatum sibiricum (P. sibiricum), recognized as a precious nourishing Chinese traditional medicine, exhibits the pharmacological effect of anti-aging. In this work, we proposed a novel mechanism underlying this effect related to the less studied bioactive compounds fructooligosaccharides in P. sibiricum (PFOS) to identify the inhibition effect of the small glycosyl molecules on the age-related zinc metalloprotease carbonic anhydrase II (CA II). Molecular docking and molecular dynamics simulation were used to investigate the structural and energetic properties of the complex systems consisting of the CA II enzyme and two possible structures of PFOS molecules (PFOS-A and PFOS-B). The binding affinity of PFOS-A (-7.27 ± 1.02 kcal/mol) and PFOS-B (-8.09 ± 1.75 kcal/mol) shows the spontaneity of the binding process and the stability of the combination in the solvent. Based on the residue energy decomposition and nonbonded interactions analysis, the C-, D- and G-sheet fragments of the CA II were found to be crucial in binding process. Van der Waals interactions form on the hydrophobic surface of CAII mainly with 131PHE and 135VAL, while hydrogen bonds form on the hydrophilic surface mainly with 67ASN and 92GLN. The binding of PFOS results in the blocking of the zinc ions pocket and then inhibiting its catalytic activity, the stability of which has been further demonstrated by free energy landscape. These findings provide evidence of the effective inhibition of PFOS to CA II enzyme, which leads to a novel direction for exploring the mechanism of traditional Chinese medicine focused on small molecule fructooligosaccharides.

Differentiation of Polygonatum Cyrtonema Hua from Different Geographical Origins by near-Infrared Spectroscopy with Chemometrics.

BACKGROUND: The identification of the geographical origin of Polygonatum cyrtonema Hua is of particular importance because the quality and market value of Polygonatum cyrtonema Hua from different production areas are highly variable due to differences in the growing environment and climatic conditions. OBJECTIVE: This study utilized near-infrared spectra (NIR) of Polygonatum cyrtonema Hua (n = 400) to develop qualitative models for effective differentiation of Polygonatum cyrtonema Hua from various regions. METHODS: The models were produced under different conditions to distinguish the origins distinctly. Ten pre-processing methods have been used to pre-process the original spectra (OS) and to select the most optimal spectral pre-processing method. Principal component analysis (PCA), partial least squares discriminant analysis (PLS-DA), and orthogonal partial least squares discriminant analysis (OPLS-DA) were employed to determine appropriate models. For simplicity, the pretreated full spectrum was calculated by different wavelength selection methods, and the four most significant variables were selected as discriminant indicator variables. RESULTS: The results show that Polygonatum cyrtonema Hua from different regions can be effectively distinguished using spectra from a series of samples analyzed by OPLS-DA. The accuracy of the OPLS-DA model is also satisfactory, with a good differentiation rate. CONCLUSION: The study findings indicate the feasibility of using spectroscopy in combination with multivariate analysis to identify the geographical origins of Polygonatum cyrtonema Hua. HIGHLIGHTS: The utilization of near-infrared spectroscopy combined with chemometrics exhibits high efficacy in discerning the provenance of herbal medicines and foods, thereby facilitating quality assurance measures.

Anti-aging mechanism and effect of treatment with raw and wine-steamed Polygonatum sibiricum on D-galactose-induced aging in mice by inhibiting oxidative stress and modulating gut microbiota.

Background: Polygonatum sibiricum (PS) is a traditional Chinese medicine (TCM) first recorded in Mingyi Bielu. The book documents that PS can nourish five internal organs, be taken for a long time, relax the body and prolong lifespan. Presently, PS is widely used in TCM to prevent premature graying of hair. Based on TCM theory and clinical trials, the wine steaming processed product from PS provides a better effect. However, no published study has elucidated the anti-aging mechanism. Purpose: The study aim was to investigate the anti-aging mechanism of PS and its wine steaming processed product in mice, specifically focusing on the effect of D-galactose (D-gal) surrounding the intestinal flora and the Kelch-like ECH-associated protein 1-nuclear factor erythroid 2-related factor 2-antioxidant response elements (Keap1/Nrf2/ARE) pathway. Methods: The chemical components in Raw PS (RPS) and Wine-steamed PS (WPS) were identified by ultra-performance liquid chromatography-hybrid quadrupole-Orbitrap high-resolution mass spectrometry (UPLC-Q-Orbitrap HRMS). An aging model using Kunming mice was established through intraperitoneally injected D-gal. Concentrations of RPS and WPS at 5, 10, or 15 g/kg/day levels were administered intragastrically, respectively. The body weight, liver and spleen indexes, superoxide dismutase (SOD), glutathione peroxidase (GSH-PX), and malondialdehyde (MDA) activities in serum and brain tissue were recorded. Hematoxylin and eosin (HE) stained brain tissue was histopathologically examined. The expressions of Keap1, Nrf2 and heme oxygenase 1 (HO-1) in the brain tissue at the mRNA and protein levels were respectively detected by reverse transcription-polymerase chain reaction (RT-PCR) and western blot (WB). Moreover, an Illumina Hiseq platform was used for 16S ribosomal RNA (16S rRNA) high-throughput sequencing to evaluate the proportions of intestinal flora in aging mice. Results: The proportions of saccharides, flavonoids, and triterpene acids were different between RPS and WPS. In the aging model mice, WPS outperformed RPS in improving body weight and mental state by increasing the spleen index, SOD and GSH-PX activities, decreasing the liver index and MDA activities, and restoring the histopathological morphology in D-gal-induced aging mice. At the mRNA levels, RPS and WPS significantly reduced the expression of Keap1 and increased the expressions of Nrf2 and HO-1. The trend in protein expressions was similar to that of the mRNA results, and WPS had a stronger effect than RPS. Fecal microbiota analysis showed that RPS and WPS restored intestinal microbiota proportions to normal levels. Conclusion: The results demonstrated that PS and its WPS had a positive effect in relieving oxidative stress in aging mice. WPS outperformed RPS, which might be related to the activation of the Keap1/Nrf2/ARE pathway and regulation of intestinal flora.

Integrated Metabolomics and Transcriptomics Analysis of Flavonoid Biosynthesis Pathway in Polygonatum cyrtonema Hua.

Flavonoids, a class of phenolic compounds, are one of the main functional components and have a wide range of molecular structures and biological activities in Polygonatum. A few of them, including homoisoflavonoids, chalcones, isoflavones, and flavones, were identified in Polygonatum and displayed a wide range of powerful biological activities, such as anti-cancer, anti-viral, and blood sugar regulation. However, few studies have systematically been published on the flavonoid biosynthesis pathway in Polygonatum cyrtonema Hua. Therefore, in the present study, a combined transcriptome and metabolome analysis was performed on the leaf, stem, rhizome, and root tissues of P. cyrtonema to uncover the synthesis pathway of flavonoids and to identify key regulatory genes. Flavonoid-targeted metabolomics detected a total of 65 active substances from four different tissues, among which 49 substances were first study to identify in Polygonatum, and 38 substances were flavonoids. A total of 19 differentially accumulated metabolites (DAMs) (five flavonols, three flavones, two dihydrochalcones, two flavanones, one flavanol, five phenylpropanoids, and one coumarin) were finally screened by KEGG enrichment analysis. Transcriptome analysis indicated that a total of 222 unigenes encoding 28 enzymes were annotated into three flavonoid biosynthesis pathways, which were "phenylpropanoid biosynthesis", "flavonoid biosynthesis", and "flavone and flavonol biosynthesis". The combined analysis of the metabolome and transcriptome revealed that 37 differentially expressed genes (DEGs) encoding 11 enzymes (C4H, PAL, 4CL, CHS, CHI, F3H, DFR, LAR, ANR, FNS, FLS) and 19 DAMs were more likely to be regulated in the flavonoid biosynthesis pathway. The expression of 11 DEGs was validated by qRT-PCR, resulting in good agreement with the RNA-Seq. Our studies provide a theoretical basis for further elucidating the flavonoid biosynthesis pathway in Polygonatum.

The structure change of polygonatum polysaccharide and the protect effect of Polygonatum crtonema Hua extracts and polysaccharide on cisplatin-induced AKI mice during nine-steam-nine-bask processing.

Polygonatum cyrtonema Hua (PC) with different processing degrees during the nine-steam-nine-bask processing was selected as the research object to investigate the changes of polysaccharide structure and their protective effect on cisplatin-induced acute kidney injury (AKI) in mice. The polysaccharides (PCP0, PCP4 and PCP9) were extracted, whose polysaccharide contents were 62.45 %, 60.34 % and 58.23 %, respectively. After processing, the apparent structure of PCPs became looser, and the apparent viscosity and the particle size were decreased. The PCPs were acidic polysaccharides containing pyran rings, and furan rings were present in PCP4 and PCP9. Besides, processing destroyed the original ß-glucoside bond in PCP0. PCPs were all composed of Rha, Man, Glu, Gal, Xyl and Ara with different ratio. In addition, AKI mice model was successfully constructed by single intraperitoneal injection of 15 mg/kg cisplatin. PC extracts (3.0750 g/kg) and PCP (0.1599 g/kg) significantly decreased the kidney function, liver function, and percentage of renal cell apoptosis, and improved the kidney structure of AKI mice (p < 0.05). PC and PCP have protective effect on cisplatin-induced AKI mice, and the protective effect was improved with the increase of processing degree. Under the same processing degree, the protective effect of PC mixed extract was better than that of PCP.

Structural Characterization of Polygonatum Cyrtonema Polysaccharide and Its Immunomodulatory Effects on Macrophages.

A neutral Polygonatum cyrtonema polysaccharide (NPCP) was isolated and purified from Polygonatum cyrtonema by various chromatographic techniques, including DEAE-52 and Sephadex-G100 chromatography. The structure of NPCP was characterized by HPLC, HPGPC, GC-MS, FT-IR, NMR, and SEM. Results showed that NPCP is composed of glucose (55.4%) and galactose (44.6%) with a molecular weight of 3.2 kDa, and the sugar chain of NPCP was →1)-α-D-Glc-(4→1)-ß-D-Gal-(3→. In vitro bioactivity experiments demonstrated that NPCP significantly enhanced macrophages proliferation and phagocytosis while inhibiting the M1 polarization induced by LPS as well as the M2 polarization induced by IL-4 and IL-13 in macrophages. Additionally, NPCP suppressed the secretion of IL-6 and TNF-α in both M1 and M2 cells but promoted the secretion of IL-10. These results suggest that NPCP could serve as an immunomodulatory agent with potential applications in anti-inflammatory therapy.

Molecular cytogenetic characterization of 9 populations of four species in the genus Polygonatum (Asparagaceae).

To characterize the chromosomes of the four species of Polygonatum Miller, 1754, used in traditional Chinese medicine, P.cyrtonema Hua, 1892, P.kingianum Collett et Hemsley, 1890, P.odoratum (Miller, 1768) Druce, 1906, and P.sibiricum Redouté, 1811, and have an insight into the karyotype variation of the genus Polygonatum, fluorescence in situ hybridization (FISH) with 5S and 45S rDNA oligonucleotide probes was applied to analyze the karyotypes of 9 populations of the four species. Detailed molecular cytogenetic karyotypes of the 9 populations were established for the first time using the dataset of chromosome measurements and FISH signals of 5S and 45S rDNA. Four karyotype asymmetry indices, CVCI, CVCL, MCA and Stebbins' category, were measured to elucidate the asymmetry of the karyotypes and karyological relationships among species. Comparison of their karyotypes revealed distinct variations in the karyotypic parameters and rDNA patterns among and within species. The basic chromosome numbers detected were x = 9, 11 and 13 for P.cyrtonema, x = 15 for P.kingianum, x = 10 and 11 for P.odoratum, and x = 12 for P.sibiricum. The original basic chromosome numbers of the four species were inferred on the basis of the data of this study and previous reports. All the 9 karyotypes were of moderate asymmetry and composed of metacentric, submetacentric and subtelocentric chromosomes or consisted of two of these types of chromosomes. Seven populations have one locus of 5S rDNA and two loci of 45S rDNA, and two populations added one 5S or 45S locus. The karyological relationships among the four species revealed by comparison of rDNA patterns and PCoA based on x, 2n, TCL, CVCI, MCA and CVCL were basically accordant with the phylogenetic relationships revealed by molecular phylogenetic studies. The mechanisms of both intra- and inter-specific dysploidy in Polygonatum were discussed based on the data of this study and literature.

Transcriptome-wide identification of ARF gene family in medicinal plant Polygonatum kingianum and expression analysis of PkARF members in different tissues.

BACKGROUND: Polygonatum kingianum holds significant importance in Traditional Chinese Medicine due to its medicinal properties, characterized by its diverse chemical constituents including polysaccharides, terpenoids, flavonoids, phenols, and phenylpropanoids. The Auxin Response Factor (ARF) is a pivotal transcription factor known for its regulatory role in both primary and secondary metabolite synthesis. However, our understanding of the ARF gene family in P. kingianum remains limited. METHODS AND RESULTS: We employed RNA-Seq to sequence three distinct tissues (leaf, root, and stem) of P. kingianum. The analysis revealed a total of 31,558 differentially expressed genes (DEGs), with 43 species of transcription factors annotated among them. Analyses via gene ontology and the Kyoto Encyclopedia of Genes and Genomes demonstrated that these DEGs were predominantly enriched in metabolic pathways and secondary metabolite biosynthesis. The proposed temporal expression analysis categorized the DEGs into nine clusters, suggesting the same expression trends that may be coordinated in multiple biological processes across the three tissues. Additionally, we conducted screening and expression pattern analysis of the ARF gene family, identifying 12 significantly expressed PkARF genes in P. kingianum roots. This discovery lays the groundwork for investigations into the role of PkARF genes in root growth, development, and secondary metabolism regulation. CONCLUSION: The obtained data and insights serve as a focal point for further research studies, centred on genetic manipulation of growth and secondary metabolism in P. kingianum. Furthermore, these findings contribute to the understanding of functional genomics in P. kingianum, offering valuable genetic resources.

Polygonatum sibiricum polysaccharide ameliorates skeletal muscle aging via mitochondria-associated membrane-mediated calcium homeostasis regulation.

BACKGROUND: Sarcopenia, an age-related disease, is characterized by a gradual loss of muscle mass, strength, and function. It has been linked to abnormal organelle function in myotubes, including the mitochondria and endoplasmic reticulum (ER). Recent studies revealed that mitochondria-associated membranes (MAM), the sites connecting mitochondria and the ER, may be implicated in skeletal muscle aging. In this arena, the potential of Polygonatum sibiricum polysaccharide (PSP) emerges as a beacon of hope. PSP, with its remarkable antioxidant and anti-senescence properties, is on the cusp of a therapeutic revolution, offering a promising strategy to mitigate the impacts of sarcopenia. PURPOSE: The objective of this research is to explore the effects of PSP on age-related muscle dysfunction and the underlying mechanisms involved both in vivo and in vitro. METHODS: In this investigation, we used in vitro experiments using D-galactose (D-gal)-induced aging in C2C12 myotubes and in vivo experiments on aged mice. Key indices were assessed, including reactive oxygen species (ROS) levels, mitochondrial function, the expression of aging-related markers, and the key proteins of mitochondria and MAM fraction. Differentially expressed genes (DEGs) related to mitochondria and ER were identified, and bioinformatic analyses were performed to explore underlying mechanisms. Muscle mass and function were determined to evaluate the quantity and quality of skeletal muscle in vivo. RESULTS: PSP treatment effectively mitigated oxidative stress and mitochondrial malfunction caused by D-gal in C2C12 myotubes, preserving mitochondrial fitness and reducing MAM formation. Besides, PSP attenuated D-gal-induced increases in Ca2+ concentrations intracellularly by modulating the calcium-related proteins, which were also confirmed by gene ontology (GO) analysis of DEGs. In aged mice, PSP increased muscle mass and improved grip strength, hanging time, and other parameters while reducing ROS levels and increasing antioxidant enzyme activities in skeletal muscle tissue. CONCLUSION: PSP offers protection against age-associated muscle impairments. The proposed mechanism suggests that modulation of calcium homeostasis via regulation of the MAM results in a favorable functional outcome during skeletal muscle aging. The results of this study highlight the prospect of PSP as a curative intervention for sarcopenia and affiliated pathological conditions, warranting further investigation.

The accumulation of active ingredients of Polygonatum cyrtonema Hua is associated with soil characteristics and bacterial community.

Introduction: With the increasing demand for health products derived from Polygonati rhizoma (PR), people begin to artificially plant Polygonatum cyrtonema Hua (P. cyrtonema) in the middle and lower reaches of the Yangtze River. To promote P. cyrtonema cultivation and increase farmers' income, efforts are needed to understand the ways to obtain high-quality PR under artificial cultivation conditions. Methods: Rhizomes of artificial planting P. cyrtonema and rhizosphere soils were collected across five regions in Zhejiang Province, China. Subsequently, the contents of the main active ingredients of P. cyrtonema and soil properties were analyzed, and both rhizosphere and endophytic bacteria of P. cyrtonema were detected by 16S rDNA sequencing. The relationship between the active ingredients and soil properties, and the dominant bacteria were investigated by correlation analysis. Results: The content of active ingredients of P. cyrtonema from the five regions varied significantly, especially polysaccharides and saponins. High-throughput sequencing demonstrated that Proteobacteria was the dominant bacterial phylum in all samples, and Burkholderia-Caballeronia-Paraburkholderia was the main endophytic bacterial genus in rhizome. In addition, the bacterial diversity and richness of rhizosphere soil samples were higher than those of rhizome samples. Soil physicochemical properties and enzyme activities were significantly different across regions, leading to notable variations in the community structures of rhizosphere and endophytic bacteria. Redundancy analysis (RDA) displayed that pH and urease (UE) were the major factors altering shifting rhizosphere bacteria community structure. Moreover, the composition and diversity of rhizome endophytic bacteria were principally affected by both soil physicochemical properties and soil enzyme activities. Soil properties and bacteria from rhizosphere soil and rhizome had a considerable impact on certain active ingredients in P. cyrtonema under artificial cultivation conditions after Pearson correlation analysis. Polysaccharides were significantly correlated with nutrient-rich soil and endophytic bacteria, such as Burkholderia-Caballeronia-Paraburkholderia, Pseudomonas, Ralstonia, and Bacillus. However, flavonoids were associated with nutrient-poor soil. Saponins were positively correlated with OM and available phosphorous (AP) and were significantly negatively affected by rhizosphere bacterial communities. Conclusion: The study demonstrated that bacterial microorganisms were involved in the accumulation of active ingredients of P. cyrtonema together with soil physicochemical properties and enzyme activities, which provided a theoretical basis for the scientific and effective artificial cultivation of high-quality P. cyrtonema.

Metabolomics and HS-SPME-GC-MS-based analysis of quality succession patterns and flavor characteristics changes during the fermentation of Lycium barbarum and Polygonatum cyrtonema compound wine.

This work set out to investigate how the physicochemical markers, volatiles, and metabolomic characteristics of mixed fermented the fermentation of Lycium barbarum and Polygonatum cyrtonema compound wine (LPCW) from S. cerevisine RW and D. hansenii AS2.45 changed over the course of fermentation. HS-SPME-GC-MS combined with non-targeted metabolomics was used to follow up and monitor the fermentation process of LPCW. In total, 43 volatile chemical substances, mostly alcohols, esters, acids, carbonyl compounds, etc., were discovered in LPCW. After 30 days of fermentation, phenylethyl alcohol had increased to 3045.83 g/mL, giving off a rose-like fresh scent. The biosynthesis of valine, leucine, and isoleucine as well as the metabolism of alanine, aspartic acid, and glutamic acid were the major routes that led to the identification of 1385 non-volatile components in total. This study offers a theoretical foundation for industrial development and advances our knowledge of the fundamental mechanism underlying flavor generation during LPCW fermentation.