Association between gut microbiota and diabetic nephropathy: a mendelian randomization study.

Background: The correlation between diabetic nephropathy (DN) and gut microbiota (GM) has been suggested in numerous animal experiments and cross-sectional studies. However, a causal association between GM and DN has not been ascertained. Methods: This research adopted MR analysis to evaluate the causal link between GM and DN derived from data acquired through publicly available genome-wide association studies (GWAS). The study utilized the inverse variance weighted (IVW) approach to assess causal association between GM and DN. Four additional methods including MR-Egger, weighted median, weighted mode, and simple mode were employed to ensure comprehensive analysis and robust results. The Cochran's Q test and the MR-Egger method were conducted to identify heterogeneity and horizontal pleiotropy, respectively. The leave-one-out approach was utilized to evaluate the stability of MR results. Finally, a reverse MR was performed to identify the reverse causal association between GM and DN. Results: According to IVW analysis, Class Verrucomicrobiae (p = 0.003), Order Verrucomicrobiales (p = 0.003), Family Verrucomicrobiaceae (p = 0.003), Genus Akkermansia (p = 0.003), Genus Catenibacterium (p = 0.031), Genus Coprococcus 1 (p = 0.022), Genus Eubacterium hallii group (p = 0.018), and Genus Marvinbryantia (p = 0.023) were associated with a higher risk of DN. On the contrary, Class Actinobacteria (p = 0.037), Group Eubacterium ventriosum group (p = 0.030), Group Ruminococcus gauvreauii group (p = 0.048), Order Lactobacillales (p = 0.045), Phylum Proteobacteria (p = 0.017) were associated with a lower risk of DN. The sensitivity analysis did not identify any substantial pleiotropy or heterogeneity in the outcomes. We found causal effects of DN on 11 GM species in the reverse MR analysis. Notably, Phylum Proteobacteria and DN are mutually causalities. Conclusion: This study identified the causal association between GM and DN with MR analysis, which may enhance the understanding of the intestinal-renal axis and provide novel potential targets for early non-invasive diagnosis and treatment of DN.

High Pressure Rapid Synthesis of LiCrTiO4 with Oxygen Vacancy for High Rate Lithium-Ion Battery Anodes.

Lithium-ion battery based on LiCrTiO4 (LCTO) is considered to be a promising anode material, as they provide higher safety and durability beyond than that of graphite electrode. However, the applications of this transformative technology demand improved inherent electrical conductivity of LCTO as well as a simple and rapid synthetic route. Here, LCTO with oxygen vacancies (OVs) is fabricated using high-pressure synthesis technology in only 40 min. The optimal synthesis pressure is 0.8 GPa (LCTO-0.8). The reversible capacity of LCTO-0.8 at 1C is 131 mA h g-1 after 1000 cycles and the capacity retention is nearly 97%, and the reversible capacity of LCTO synthesized at atmospheric pressure (LCTO-P) is 85 mA h g-1 under the same circumstances. Even at 5C, the reversible capacity is 110 mA h g-1 , which is 77% higher than LCTO-P. Furthermore, it is confirmed by theoretical calculations that the introduction of OVs has the occupation of electronic states at the Fermi level, which greatly enhances the intrinsic conductivity of LCTO. Specifically, the electronic conductivity has increased by two orders of magnitude compared with LCTO-P. Therefore, high-pressure synthesis technology endows LCTO with superior characteristics, providing a new avenue for industrialization.

Niclosamide attenuates inflammatory cytokines via the autophagy pathway leading to improved outcomes in renal ischemia/reperfusion injury.

Renal ischemia/reperfusion (I/R) injury is a debilitating condition that leads to loss renal function and damage to kidney tissue in the majority of patients with acute kidney disease. Previous studies have indicated that autophagy serves a protective function in renal I/R injury. In the present study, the effect of the anthelmintic niclosamide in the regulation of inflammatory responses in kidney I/R was investigated. A total of 40 Sprague-Dawley rats were randomly divided into the following 5 groups (n=8 in each group): Sham group; renal I/R injury; renal I/R injury plus 3­methyladenine (3­MA) treatment (15 mg/kg); renal I/R injury plus niclosamide (25 mg/kg); and renal I/R injury plus rapamycin (10 mg/kg). The expression levels of autophagy­associated proteins in kidney samples obtained from rats with I/R injury were examined using reverse transcription­quantitative polymerase chain reaction and western blotting techniques. In addition, histopathological alterations, the expression of cytokines and renal function were evaluated. Treatment with niclosamide was associated with induction of autophagy and an overall improvement in renal function. There was an increased expression of autophagosome­associated proteins, suggesting a strong correlation between autophagy and improvement of renal function. The increased levels of anti­inflammatory cytokines and decreased levels of pro­inflammatory cytokines provided additional evidence that niclosamide may be effective for the treatment of renal I/R injury. Clinical studies are required to further validate the results of the present study.

[Preparation Technique of Tetraploid of Dioscorea zingiberensis Synthetic Seed Based on Embryogenic Callus].

OBJECTIVE: To provide technical support for industrialization promotion of tetraploid of Dioscorea zingiberensis, the manufacturing method for synthetic seeds of tetraploid of Dioscorea zingiberensis was established and the correlated influential factors were studied. METHODS: By taking embryogenic calluses of tetraploid of Dioscorea zingiberensis as propagation materials, the influential factors such as components of artificial endosperm, seed coats,storage conditions and germination materials on germination and seedling of the synthetic seeds were evaluated. RESULTS: When 4% alginate +2% CaCl2 + 2% chitosan was served as seed coat materials, and 1/2 MS +0. 2 mg/L BA +0. 5 mg/L NAA + 0. 1 mg/L penicillin + 0. 3% carhendazim powder + 0. 2% sodium benzoate + 1. 0% sucrose + 0. 5% activated carbon + 1. 0% tapioca starch was served as endosperm, the synthetic seeds had high germination rate and seedling rate. After storing at 4 °C for 20 d, the germination rate and seedling rate of synthetic seeds was 76. 7% and 71. 7%, respectively. CONCLUSION: Manufacturing technology of synthetic seeds of tetraploid of Dioscorea zingiberensis with embryogenic calluses as propagation materials has production prospects.

[Genetic stability study on autotetraploid plant of Dioscorea zingiberensis].

OBJECTIVE: To study the genetic stability of autotetraploid plant of Dioscorea zingiberensis. METHODS: The chromosome of root-tip was determined by photomicroscope, and the agronomic characters were observed in the period of stable growth. The protein content was determined and the experiment of protein polyacrylamide gel electrophoresis was carried out. Furthemore, the diosgenin content was determined and compared. RESULTS: The chromosome number of autotetraploid plantlet was 2n = 4x = 40. The agronomic characters showed typical autotetraploid characteristics. The contents of diosgenin and protein of autotetraploid were higher than that of the diploid. The protein electrophoresis bands of all the lines were similar. CONCLUSION: The experiment confirmed that the autotetraploid plant of Dioscorea zingiberensis, which was artificially induced, had good genetic stability. It lays the foundation for the polyploid breeding to develop superior varieties of Dioscorea zingiberensis.

Germplasm preservation in vitro of Polygonum multiflorum Thunb.

BACKGROUND: The root of Polygonum multiflorum Thunb. is a common traditional Chinese medicine. In recent years, the wild resources of P. multiflorum have been seriously broken, and the cultivated varieties have been degrading. The germplasm resources of P. multiflorum need protection and preservation. So far, no in vitro germplasm preservation of P. multiflorum has been reported. OBJECTIVE: To explore a method for the in vitro germplasm preservation of P. multiflorum. MATERIALS AND METHODS: A large number of buds from seed explants were induced by tissue culture. The single buds were used as experimental materials to study the effects of plant growth regulator, temperature, and osmotic pressure on the preservation time, growth recovery, and genetic stability. RESULTS: When the buds were inoculated onto Murashige and Skoog (MS) basal media containing 4% w/v sucrose, 2% w/v mannitol, and 1% w/v sorbitol, supplemented with paclobutrazol (PP333) 1.0 mg/l, abscisic acid (ABA) 5.0 mg/l, and daminozide (B9) 30.0 mg/l in an illuminated chamber under a 16 h photoperiod of 1500 lx light intensity at 15°C for 10 months, the survival rate was over 70% with good growth recovery and genetic stability. CONCLUSION: The results of this study can be used for medium-term in vitro germplasm preservation of P. multiflorum, and meeting actual needs of research and production.

Autotetraploidy induced in nianmaohuangqin (Radix Scutellariae Viscidulae) with colchicine in vitro.

OBJECTIVE: To establish and optimize the propagation of Nianmaohuangqin (Radix Scutellariae Viscidulae) and induce and characterize polyploidy of Nianmaohuangqin (Radix Scutellariae Viscidulae). METHODS: Buds from germinating seed-derived explants were induced by tissue culture. With an orthogonal test, different concentrations of 6-benzyl-aminopurine (BAP), indole-3-acetic acid (IAA) and kinetin (KT) were used to determine the optimal concentrations for the propagation of Nianmaohuangqin (Radix Scutellariae Viscidulae). The different concentrations of IAA and rooting powder (ABT) were used to induce rooting. A 0.3% w/v colchicine solution was used to induce polyploidy and the induced buds was identified by root-tip chromosome determination and stomatal apparatus observation. RESULTS: A large number of buds could be induced directly from epicotyl and hypocotyl explants on Murashige and Skoog (MS) (Murashige and Skoog 1962) medium supplemented with 1.1-1.3 mg/L BAP and 0.2 mg/L IAA. Root induction and development could be observed within 20 days of inoculation on 1/2 MS medium supplemented with 0.2 mg/L IAA and 0.1 mg/L ABT. Furthermore, 27 lines of autotetraploid individuals were obtained with a plantlet chromosome number of 2n = 4x = 36. CONCLUSION: Autotetraploid lines could be obtained through induction with colchicine in vitro, proving that this method might be used for plant selection and breeding.

[Study on inducing embryogenic callus of Dioscorea zingiberensis by orthogonal design].

OBJECTIVE: To select the suitable medium to induce embryogenic callus of Dioscorea zingiberensis. METHODS: Plantlet of Dioscorea zingiberesis in vitro was obtained by using apical meristem as explant. The different parts of the plantlets were cultured to select the best explant used for inducing callus and embryoids. Growing rate and diosgenin content were calculated in orthogonal test to optimize combination of phytohormones for inducing embryogenic callus. RESULTS: The leaves were suitable explants to induce callus and embryoid. The inducing rate of callus and embryoids reached 92.5% and 42.5%, respectively. The optimal medium for inducing embryogenic callus was MS + 6-BA 2.0 mg/L + NAA 0.5 mg/L + 2,4-D 1.0 mg/L. CONCLUSION: The results of this study can be used for effective induction of embryogenic callus of Dioscorea zingiberensis, and lay the foundation for the subsequent research of artificial seeds.

[Optinization of rapid propagation technique and induction and identification of autotetraploid of Polygonum multiflorum].

OBJECTIVE: To establish and optimize the rapid propagation system of Polygonum multiflorum, as well as explore method for induction and identification of autotetraploid. METHOD: Propagation medium was optimized by orthogonal test. The buds were immersed in colchicine solution with different concentrations for different time to select induction conditions for autotetraploid of P. multiflorum. RESULT: The most appropriate propagation medium was MS medium supplemented with 1.0 mg x L(-1) 6-BA, 0.3 mg x L(-1) NAA, and 0.4 mg x L(-1) PP333. That the buds were soaked in 0.2% colchicine solution for 30 h, or soaked in 0.3% colchicine solution for 18 h, was optimal condition to induce autopolyploid of P. multiflorum with induction rate as high as 16.7%. CONCLUSION: Rapid propagation of P. multiflorum could be achieved by tissue culture. Furthermore, colchicine was an effective inducer of polyploidy, and 25 tetraploid lines were obtained through chromosome identification. The experiment laid a foundation for the wild resource conservation, superior varieties breeding of P. multiflorum.

[Studies on adventitious root induction in vitro and suspension culture of Polygonum multiflorum].

To achieve sustainable resources use of Polygonum multiflorum, adventitious roots were efficiently induced and cultured by suspension culture. In order to obtain optimal medium for induction adventitious roots from the young stems of P. multiflorum, MS medium was optimized by supplementing with different concentrations of sucrose and plant growth substances. The optimal medium for suspension culture of adventitious roots was determined by orthogonal design. The adventitious roots with suspension culture were subcultured, and the growth curve was also determined. Furthermore, the effective compound in adventitious roots was detected. The result indicated that the optimal medium for efficient induction of adventitious roots was MS medium containing 4% w/v sucrose, supplemented with 2.0 mg x L(-1) NAA, and 0.2 mg x L(-1) 6-BA. The optimal medium for suspension culture of adventitious roots was MS medium containing 3% sucrose, supplemented with 2.0 mg x L(-1) NAA, and 0.2 mg x L(-1) ABT-7.2,3,5,4'-tetrahydroxyl-diphenyl-ethylene-2-O-beta-D-glucoside was detected in adventitious roots, which was effective compound in medicinal material of P. multiflorum. In conclusion, the experiment achieved efficient induction and suspension culture of adventitious roots of P. multiflorum, and laid a foundation for the research on the sustainable use of traditional Chinese medicine resources.

[Effects of 1, 25(OH)(2);D(3); on parathyroid hormone induced transdifferentiation and TGF-ß(1); expression in cultured human renal tubular epithelial cells].

AIM: To explore the effects of 1, 25(OH)(2);D(3); on parathyroid hormone (PTH) induced transdifferentiation and TGF-ß(1); expression in cultured human renal tubular epithelial cells. METHODS: HK-2 cells were cultured in DMEM/F12 medium supplemented with 50 mL/L FBS. Cells were divided into three groups. (1) CONTROL GROUP: without PTH or 1, 25(OH)(2);D(3);; (2) PTH group: 10(-10); mol/L PTH; (3) PTH and 1, 25(OH)(2);D(3); group: 10(-10); mol/L PTH and different concentrations of 1, 25(OH)(2);D(3); (10(-10);, 10(-9);, 10(-8); and 10(-7); mol/L). The gene expressions of α-SMA and TGF-ß(1); were detected by semi-quantitative RT-PCR. The protein expressions of α-SMA and TGF-ß(1); were detected by Western blot. Immunocytochemisty (ICC) was used to measure the expression of α-SMA in HK-2. ELISA was used to assay the level of TGF-ß(1); in the supernatant. RESULTS: The gene expressions of α-SMA and TGF-ß(1); in PTH group were significantly higher than those in control group (P<0.05). In contrast, they were significantly lower in PTH and 1, 25(OH)(2);D(3); group than those in PTH group (P<0.05). Western blot results showed α-SMA could not be detected in normal HK-2 cells, which could be detected in PTH group. TGF-ß(1); protein expression in PTH group was higher than that in control group. In PTH and 1, 25(OH)(2);D(3); group, α-SMA and TGF-ß(1); protein expressions were significantly lower than those in PTH group (P<0.05). ICC results showed that α-SMA was hardly expressed in cells of control group. However, positive expression of α-SMA could be seen in many cells in PTH group. In PTH and 1, 25(OH)(2);D(3); group, the cells of α-SMA positive expressed were significantly less than those in PTH group (P<0.05). ELISA results showed that the level of TGF-ß(1); in the supernatant of PTH group was higher than that in control group, which was also higher than that in PTH and 1, 25(OH)(2);D(3); group (P<0.05). CONCLUSION: 1, 25(OH)(2);D(3); can attenuate PTH-induced transdifferentiation and TGF-ß(1); expression in cultured human renal tubular epithelial cells.

Up-regulation of licochalcone A biosynthesis and secretion by Tween 80 in hairy root cultures of Glycyrrhiza uralensis Fisch.

We evaluated the effect of Tween 80 as elicitor on licochalcone A from hairy root cultures of Glycyrrhiza uralensis Fisch. After a 15-days treatment with 2% Tween 80, hairy roots still grew well and produced higher levels of licochalcone A and total flavonoids than the control (without treatment). Licochalcone A content and total flavonoid content were 3.103 and 127.095 mg per flask (9- and 11-fold higher), respectively, compared with controls. Secretion of licochalcone A and total flavonoids into the culture medium was remarkably high, up to 98 and 94% of the total production, respectively. The enhanced flavonoid production was associated with elevated mRNA levels and enzyme activities of phenylalanine ammonia-lyase (PAL), 4-coumarate:coenzyme A ligase (4CL), and cinnamate-4-hydroxylase (C4H). These results clearly demonstrated that Tween 80 treatment permeabilized the roots to enhance secretion, but also acted as an efficient elicitor of licochalcone A and total flavonoid production in hairy roots of G. uralensis Fisch.

In vitro tetraploid induction and generation of tetraploids from mixoploids in Dioscorea zingiberensis.

This article describes an efficient colchicine-mediated technique for the in vitro induction of tetraploids in Dioscorea zingiberensis and its confirmation by flow cytometry. Buds immersed in 0.2% colchicine solution for 36 hours prior to culture induced as high as 35.6% tetraploid plants. Colchicine-induced tetraploids remained stable after six months in soil. Leaf characteristics of diploids and tetraploids in D. zingiberensis were compared. It was determined that the leaf sizes of glasshouse-grown plants and stomatal sizes of both in vitro and glasshouse-grown plants were suitable parameters for identifying putative tetraploids in D. zingiberensis. Besides generating tetraploids, this technique generated mixoploids in D. zingiberensis. Calli derived from mixoploid leaves were induced to form buds and shoots. Individual shoots were classed as diploid, mixoploid, and tetraploid by flow cytometry. This callus-based technique could be employed when a genome-doubling agent generated mixoploids, but no tetraploids.

[Culture of transgenic Glycyrrhiza uralensis hairy root with licorice squalene synthase (SQS) gene].

A squalene synthase gene cloned (GuSQS1, accession number in GenBank database: AM182329) from Glycyrrhiza uralensis was transferred into G. uralensis via Agrobacterium rhizogenes A4 for investigating biosynthesis pathway and enhancing synthesis of glycyrrhizic acid (GA). Hypocotyl explants from G. uralensis were infected with A. rhizogenes A4 containing GuSQS1 gene to induce the hairy roots. The hairy root lines established were selected in medium containing 0.8 mg x L(-1) phosphinothricin (PPT) and analyzed by PCR and southern blotting. The transgenic hairy roots were cultured in liquid MS medium. GA contents in transgenic hairy roots were detected by HPLC. Results showed that maximal GA content in transgenic hairy root lines was 3.6 times as high as in wild type hairy roots.

[Analysis of biochemical indices and efficient ingredients in autotetraploid of Dioscorea zingibiernsis].

OBJECTIVE: To improve the yield and quality of Dioscrea zingibiernsis. METHODS: The Biochemical indices and the diosgenin's content were analysed in the autotetraploid lines. RESULTS: The results showed that the activities of APX, SOD and POD in most of autotetraploid lines were higher than that in diploid line or close to it, there was also difference between autotetraploid lines and control lines in the SDS-PAGE of soluble proteins. The content of diosgenin in most autotetraploid plantlets were higher than that in the control. CONCLUSION: There were difference between autotetraploid and control lines in the content of diosgenin and Biochemical indices, therefore, inducing autotetraploid could be an effective way to breeding the superior varieties.

Enhanced flavonoid production in hairy root cultures of Glycyrrhiza uralensis Fisch by combining the over-expression of chalcone isomerase gene with the elicitation treatment.

Economically important compounds, such as licorice flavonoids, are present in insufficient amounts in the hairy roots. To overcome this problem, we took the transgenic approach combined with the elicitation technique to increase the flavonoid production. The Glycyrrhiza uralensis Fisch cDNA encoding chalcone isomerase gene (chi) was over-expressed in hairy roots of G. uralensis Fisch mediated by the disarmed Agrobacterium rhizogenes A4. Stable genetic transformation was confirmed by Southern blot analysis. The transgenic and wild cultures were subsequently elicited with PEG8000 (2%) alone, yeast extract (YE) (0.1%) alone, or both of them, and then the total flavonoids were extracted and measured. The results showed that over a culture period of 3 weeks, the wild-type hairy roots, the untreated transgenic hairy roots, and the double-treated transgenic hairy roots accumulated 0.842, 1.394, and 2.838 (g/100 g DW) of total flavonoids, respectively. Moreover, the enhanced accumulation of flavonoids were correlated with the elevated level of chi transcripts and CHI activity, confirming the key role of chi in the flavonoids synthesis. This research demonstrated that the combination of the metabolic engineering and PEG8000-YE elicitation treatment was an effective strategy to increase the flavonoids production in hairy roots of G. uralensis Fisch.

[Application and study of HPLC fingerprint of Lonicera japonica for identification of geo-authenticity].

OBJECTIVE: To establish a chromatographic fingerprint of Lonicera japonica and apply it to identify geo-authentic of L. japonica. METHOD: The HPLC was applied in chromatographic separation and data were analysed by "computed aided similarity evaluation" software. RESULT: There exited distinct difference of chemical components group between and non-authentic samples. CONCLUSION: The established HPLC fingerprint can be used for the identification of geo-authentic of L. japonica.

[Induction and identification of autotetraploid plant of Platycodon grandiflorum].

The methods and technique of inducing polyploid plants of Platycodon grandiflorum were done. Many autotetraploid plant lines were induced by colchicine in vitro successfully. The identification of main agronomic characteristics of these clonal lines was conduced in the fields. The results showed the giant type occurred in tetraploid plants comparing with that in diploid plants. It may show prospective way in developing new varieties with high yield and high content of useful compounds.