OBJECTIVETo investigate the antioxidant and cytotoxic properties of five diarylheptanoids (1-5) isolated from the rhizomes of Zingiber officinale.

METHODVarious models such as scavenging superoxide anions and 1,1-diphenyl-2- picrylhydrazyl (DPPH) radicals, inhibiting lipid peroxidation, as well as protecting of rat pheochromocytoma (PC12) cells induced by hydrogen peroxide (H2O2) were employed to assay the antioxidative effects of the diarylheptanoids. The cytotoxicities of compounds 1-5 were measured with MTT assays.

RESULTThe test compounds (1-5) showed promising DPPH inhibitory activities, and compound 5 exhibited the strongest DPPH scavenging activity with an IC50 value of (22.6+/-2.4) micromol x L(-1). Compounds 1, 3 and 4 showed potential anti-peroxidative effects with inhibitory rates of (66.3+/-15.4)%, (68.7+/-15.8)% and (72.2+/-10.6)%, respectively, at 100 microg x mL(-1). It could be observed that compounds 1, 3 and 4 demonstrated significant neuroprotective activities in a dose-dependent manner. Moreover, compound 3 exhibited certain cytotoxicities against human chronic myelogenous leukemia cells (K562) and its adriamycin-resistant cells (K562/ADR) with IC50 values of (34.9+/-0.6), (50.6+/-23.5) micromol x L(-1), respectively.

CONCLUSIONIn vitro results demonstrated that five diarylheptanoids (1-5) isolated from the roots of Z. officinale were capable of scavenging radicals, inhibiting lipid peroxidation and protecting PC12 cells against the insult by H2O2. Additionally, compound 3 could inhibit the growth of K562 and K562/ADR cells.

Animals ; Antioxidants ; toxicity ; Cell Proliferation ; drug effects ; Cytotoxins ; toxicity ; Diarylheptanoids ; isolation & purification ; metabolism ; toxicity ; Free Radicals ; metabolism ; Ginger ; chemistry ; Humans ; Hydrogen Peroxide ; metabolism ; K562 Cells ; Oils, Volatile ; pharmacology ; PC12 Cells ; Rats ; Rats, Sprague-Dawley

Objective:To evaluate the diagnostic value of metagenomic next-generation sequencing (mNGS) in tuberculous meningitis (TBM).Methods:From August 1, 2020 to August 31, 2022, 99 patients with suspected TBM admitted to the Tuberculosis Diagnosis and Treatment Center, Hangzhou Chest Hospital, Zhejiang University School of Medicine were enrolled. The cerebrospinal fluid (CSF) was tested for mNGS, GeneXpert Mycobacterium tuberculosis/rifampin (GeneXpert MTB/RIF) and mycobacterium culture. The sensitivity, specificity, positive predictive value, negative predictive value, agreement rate, Kappa value of the diagnostic efficacy of the three test methods were compared.The receiver-operating characteristic (ROC) curve of the diagnostic efficacy of mNGS was drawn. Chi-square test and rank sum test were used for statistical analysis. Results:Among the 99 suspected patients with TBM, 67 were diagnosed with TBM and 32 were non-TBM. Based on the results of 67 cases clinically diagnosed with TBM, the sensitivity, specificity, positive predictive value, negative predictive value, agreement rate and Kappa value of mNGS for the diagnosis of TBM were 82.1%, 100.0%, 100.0%, 72.7%, 87.9% and 0.748, respectively. The sensitivity of mNGS was higher than that of GeneXpert MTB/RIF (50.7%) and mycobacterium culture (28.4%). The differences were statistically significant ( χ2=12.61 and 32.42, respectively, both P<0.01). The detection time of mNGS was 1.0 (1.0, 2.0) day, which was shorter than 42.0 (42.0, 42.0) days of mycobacterium culture with statistical significance ( Z=10.80, P<0.001). ROC curve analysis showed that mNGS had the best diagnostic efficacy when the number of Mycobacterium tuberculosis sequences in CSF was one. Conclusions:The sensitivity and specificity of mNGS in the diagnosis of TBM are high, and the detection time is shorter, which could be used in the early diagnosis of TBM.

【Objective】  To study the effects of soil environment on the growth, yield, and quality of Sharen (Amomi Fructus) under different planting patterns. 【Methods】  Soil physical and chemical indices and enzyme activities in four periods including early flowering (March), full flowering (June), fruit ripening (September), and late fruit picking (December), were measured under three planting patterns including natural forest, greenhouse, and rubber forest in Xishuangbanna, China. The changes in soil indices during the growth periods of Sharen (Amomi Fructus) under different planting patterns were analyzed, and the differences in plant growth, yield, and quality under different planting patterns were explored. Pearson correlation analysis was used to analyze the relationship between soil indices and Sharen (Amomi Fructus) growth, yield, and quality. Principal component analysis was used to investigate the effects of soil environment under different planting patterns on Sharen (Amomi Fructus) growth, yield, and quality. 【Results】  The soil moisture, available potassium content, and urease activity of the three planting patterns of Sharen (Amomi Fructus) increased initially and decreased afterwards throughout the year; pH and organic matter content showed little change in the whole year. Exchangeable manganese content and acid phosphatase activity gradually increased throughout the year. Hydrolyzed nitrogen content, exchangeable calcium content, available zinc content, protease activity, and sucrase activity decreased initially and increased afterwards throughout the year. Exchangeable magnesium content, available iron content, and catalase activity decreased annually. Total nitrogen content, total phosphorus content, and available phosphorus content fluctuated throughout the year. The total potassium content under natural forest and greenhouse planting decreased throughout the year, while the total potassium content under rubber forest showed an upward trend all year round. The organic matter content, total nitrogen content, total potassium content, available potassium content, available zinc content, urease activity, acid phosphatase activity, and catalase activity under greenhouse were significantly lower than those under natural and rubber forests (P < 0.05). Correlation analysis showed that plant growth, yield, and quality of Sharen (Amomi Fructus) were significantly correlated with soil organic matter, total nitrogen, hydrolyzed nitrogen, total phosphorus, available phosphorus, total potassium, available potassium, exchangeable manganese, exchangeable magnesium, exchangeable calcium, available zinc, urease, acid phosphatase, and invertase (P <  0.05). The results of the principal component analysis indicated that the soil environment of Sharen (Amomi Fructus) under natural forest was the best, followed by rubber forest and greenhouse. The order of its advantages and disadvantages is consistent with the growth index of Sharen (Amomi Fructus), but contrary to the yield of Sharen (Amomi Fructus), indicating that the soil environment directly affects the growth index and nutritional components of plants. 【Conclusion】  Different planting patterns of Sharen (Amomi Fructus) have different soil nutrient content, and the change rules in the growths period are not similar, with some differences. Soil indices have impacts on plant growth, yield, and quality of Sharen (Amomi Fructus). Soil ecological environment is positively correlated with the growth characteristics of Sharen (Amomi Fructus) plants, but has no direct correlation with yield and quality.

ObjectiveTo screen out the transcriptomes related to the intervention of Wuzi Yanzongwan on the spermatogenic function of semi-castrated male mice， and to explore its potential mechanism in the intervention of the progress of low spermatogenic function. MethodBalb/c mice were randomly divided into sham-operated group， model group， testosterone propionate group（0.2 mg·kg^-1·d^-1， intramuscular injection） and Wuzi Yanzongwan group（1.56 g·kg^-1·d^-1， intragastric administration） according to body weight， with 12 mice in each group. The right testicle and epididymis were extracted from the model group and the drug administration group to construct the semi-castrated model of low spermatogenic function， while the fur and the right scrotum of the sham-operated group were only cut and immediately sterilized and sutured. At the end of the intervention， hematoxylin-eosin（HE） staining was used to observe the histopathology of testis， enzyme-linked immunosorbent assay（ELISA） was used to detect the levels of serum testosterone（T）， luteinizing hormone（LH） and follicle stimulating hormone（FSH）. The sperm count and motility of epididymis were measured by automatic sperm detector of small animal. Transcriptomic microarray technology was used to detect the mRNA expression level of testicular tissue in each group， the transcriptome of genes related to the regulation of Wuzi Yanzongwan was screened， and three mRNAs were selected for Real-time fluorescence quantitative polymerase chain reaction（Real-time PCR） to verify the transcriptome data. Through the annotation analysis of Gene Ontology（GO） and the signaling pathway analysis of Kyoto Encyclopedia of Genes and Genomes（KEGG）， the related functions of drugs regulating transcriptome were analyzed. ResultCompared with the sham-operated group， the testicular tissue of mice in the model group showed spermatogenic injury， contraction and vacuolization of the seminiferous tubules， reduction of spermatogenic cells at all levels， widening of the interstitial space， obstruction of spermatogonial cell development and other morphological abnormalities， and serum T significantly decreased， LH significantly increased（P<0.01）， and FSH elevated but no statistically significant difference， the count and vitality of epididymal sperm significantly decreased（P<0.01）. There were 882 differentially expressed mRNAs in the testicular tissues， of which 565 were up-regulated and 317 were down-regulated. Cluster analysis showed that these differentially expressed mRNA could effectively distinguish between the sham-operated group and the model group. Compared with the model group， the damage to testicular tissue in the Wuzi Yanzongwan group was reduced， the structure of the seminiferous tubules was intact， vacuolization was reduced， and the number of spermatogenic cells at all levels was significantly increased and arranged tightly. The serum T significantly increased， LH significantly decreased（P<0.01）， and FSH decreased but the difference was not statistically significant. The count and vitality of sperm in the epididymis were significantly increased（P<0.01）. Moreover， Wuzi Yanzongwan could regulate 159 mRNA levels in the testes of semi-castrated mice， of which 32 were up-regulated and 127 were down-regulated， and the data of the transcriptome assay was verified to be reliable by Real-time PCR. GO and KEGG analysis showed that the transcriptome functions regulated by Wuzi Yanzongwan were involved in the whole cell cycle process of sperm development such as sex hormone production of interstitial cells in testis， renewal， differentiation， metabolism， apoptosis and signal transduction of spermatogenic cells， and were closely related to the biological behaviors of signaling pathways such as spermatogenic stem cell function， endoplasmic reticulum protein processing and metabolic program. ConclusionWuzi Yanzongwan can effectively improve the low spermatogenic function of semi-castrated male mice， and its mechanism may be related to the regulation of testicular transcriptional regulatory network， the synthesis of sex hormones in testicular interstitial cells， the function of spermatogenic stem cells， the whole cell cycle process of spermatogenesis， as well as the expression of endoplasmic reticulum protein processing and metabolic program related genes transcription.

We explored the improved method to prepare decellularized kidney scaffold and provide experimental basis for kidney tissue engineering and renal pathology and toxicology in vitro research. We perfused rat kidneys with PBS (group control) and prepared the decellularized kidney scaffolds with sodium dodecyl sulfate (SDS) (group S), Triton X-100 combined with SDS (group TS), and Triton X-100 combined with SDS after repeated freezing and thawing (group FTS) in different flow velocity. Meanwhile we measured their fluid distributions and vascular resistance. We examined the degree of decellularization of acellular scaffolds by HE, DAPI staining and DNA quantification. We examined the retention of main composition and structural integrity of decellularized scaffolds by Masson, PAS and immunohistochemical staining. We also detected the ultrastructure, cytotoxicity and the level of growth factor of the scaffolds by scanning electron microscope, MTT and ELISA, respectively. The results showed that the time of decellularization in group FTS was less than that in group S and TS. The vascular resistance of scaffolds decellularized at 10 mL/min flow velocity was lower. The fluid distribution in groups S, TS and FTS was different from that in control group. No residual cell was detected by HE and DAPI staining. DNA content was less than 50 ng/mg. Masson, PAS and immunohistochemical staining results showed that there was extracellular collagen, polysaccharide, type I collagen, type IV collagen, fibronectin and laminin in the decellularized scaffolds, and the scanning electron microscope result showed the scaffolds had the honeycomb structure. The cytotoxicity level of decellularized scaffolds was between grade 0 to 1. The level of VEGF, EGF, IGF-1 and PDGF-BB in group FTS were significantly higher than those in group S and TS. In concluding, combining freeze-thawing with perfusion can produce more ideal and effective whole organ decellularized scaffold of rat kidney, and make a foundation for the study of kidney tissue engineering and in vitro pathology and toxicology of kidney.
Animals ; Collagen ; Extracellular Matrix ; Freezing ; Kidney ; Perfusion ; Rats ; Tissue Engineering ; Tissue Scaffolds

Metastasis is crucial for the mortality of non-small cell lung carcinoma (NSCLC) patients. The epithelial-mesenchymal transition (EMT) plays a critical role in regulating tumor metastasis. Glioma-associated oncogene 1 (Gli1) is aberrantly active in a series of tumor tissues. However, the molecular regulatory relationships between Gli1 and NSCLC metastasis have not yet been identified. Herein, we reported Gli1 promoted NSCLC metastasis. High Gli1 expression was associated with poor survival of NSCLC patients. Ectopic expression of Gli1 in low metastatic A549 and NCI-H460 cells enhanced their migration, invasion abilities and facilitated EMT process, whereas knock-down of Gli1 in high metastatic NCI-H1299 and NCI-H1703 cells showed an opposite effect. Notably, Gli1 overexpression accelerated the lung and liver metastasis of NSCLC in the intravenously injected metastasis model. Further research showed that Gli1 positively regulated Snail expression by binding to its promoter and enhancing its protein stability, thereby facilitating the migration, invasion and EMT of NSCLC. In addition, administration of GANT-61, a Gli1 inhibitor, obviously suppressed the metastasis of NSCLC. Collectively, our study reveals that Gli1 is a critical regulator for NSCLC metastasis and suggests that targeting Gli1 is a prospective therapy strategy for metastatic NSCLC.