[Effect and mechanism of Jiedu Huoxue Decoction in regulating YAP/ACSL4 pathway to inhibit ferroptosis in treatment of acute kidney injury].

Jiedu Huoxue Decoction(JDHX), first recorded in the Correction on Errors in Medical Works by WANG Qing-ren, is an effective formula screened out from ancient formulas by the traditional Chinese medicine(TCM) master ZHANG Qi to treat acute kidney injury(AKI) caused by heat, toxicity, stasis, and stagnation. This paper elucidated the therapeutic effect of JDHX on AKI and probed into the potential mechanism from ferroptosis. Thirty-two male C57BL/6 mice were randomized into four groups(n=8): normal, model, and low-and high-dose JDHX. Since the clinical treatment of AKI depends on supportive or alternative therapies and there is no specific drug, this study did not include a positive drug group. The low dose of JDHX corresponded to half of clinically equivalent dose, while the high dose corresponded to the clinically equivalent dose. Mice were administrated with JDHX by gavage daily for 7 consecutive days, while those in the normal group and the model group were administered with the corresponding volume of distilled water. On day 5 of drug administration, mice in other groups except the normal group were injected intraperitoneally with cisplatin solution at a dose of 20 mg·kg~(-1) to induce AKI, and the normal group was injected with saline. All of the mice were sacrificed 72 h after modeling, blood and kidney samples were collected for subsequent analysis. The levels of serum creatine(Scr) and blood urea nitrogen(BUN) were measured by the commercial kits. The expression level of kidney injury molecule 1(KIM-1) in the serum was measured by enzyme-linked immunosorbent assay. Hematoxylin-eosin(HE) staining, periodic acid-Schiff(PAS) staining, and Prussian blue staining were employed to observe the pathological changes, glycogen deposition, and iron deposition, respectively, in the renal tissue. In addition, the levels of glutathione(GSH), superoxide dismutase(SOD), and catalase(CAT) in the renal tissue were examined by biochemical colorimetry. Western blot was performed to determine the protein levels of acyl-CoA synthetase long chain family member 4(ACSL4), lysophosphatidylcholine acyltransferase 3(LPCAT3), and Yes-associated protein(YAP, a key molecule in the Hippo pathway) in the renal tissue. Immunohistochemistry was then employed to detect the location and expression of YAP in the renal tissue. Real-time fluorescence quantitative polymerase chain reaction(qRT-PCR) was performed to measure the mRNA levels of ACSL4 and glutathione peroxidase 4(GPX4). Compared with the normal group, the model group showed elevated serum levels of Scr, BUN, and KIM-1. In the AKI model group, the tubular epithelial cells underwent atrophy and necrotic detachment, disappearance of brush border, and some tubules became protein tubules or experienced vacuole-like degeneration. In addition, this group presented widening of the interstitium or even edema, increased renal tubule injury score, and obvious glycogen and iron deposition in parts of the renal tissue. Moreover, the model group had lower GSH, SOD, and CAT levels, higher ASCL4 and LPCAT3 levels, and lower GPX4 expression and higher YAP expression than the normal group. Compared with the model group, high dose of JDHX effectively protected renal function, lowered the levels of Scr, BUN and KIM-1, alleviated renal pathological injury, reduced glycogen and iron deposition, and elevated the GSH, SOD, and CAT levels in the renal tissue. Furthermore, JDHX down-regulated the protein levels of ACSL4, LPCAT3, and YAP and up-regulated the level of GPX4, compared with the model group. In conclusion, JDHX can protect mice from cisplatin-induced AKI by inhibiting ferroptosis via regulating the YAP/ACSL4 signaling pathway.

Research progress on the treatment of diabetic nephropathy with leech and its active ingredients.

Diabetic nephropathy (DN) is a major microvascular complication of diabetes and a common cause of chronic kidney disease. There is currently a lack of effective treatments for DN, and the prognosis for patients remains poor. Hirudin, one of the primary active components derived from leeches, demonstrates anti-coagulant, anti-fibrotic, anti-thrombotic, and anti-inflammatory properties, exhibiting significant protective effects on the kidneys. In recent years, there has been a surge of interest in studying the potential benefits of hirudin, especially in its role in the management of DN. This article delves into the mechanisms by which hirudin contributes to the treatment of DN and its clinical efficacy.

Transcriptomic analysis of Cucumis sativus uncovers putative genes related to hormone signaling under low temperature (LT) and high humidity (HH) stress.

Climate change has caused changes in environmental conditions, leading to both low temperature (LT) and high humidity (HH) stress on crops worldwide. Therefore, there is a growing need to enhance our understanding of the physiological and molecular mechanisms underlying LT and HH stress tolerance in cucumbers, given the significance of climate change. The findings of this study offer a comprehensive understanding of how the transcriptome and hormone profiles of cucumbers respond to LT and HH stress. In this study, cucumber seedlings were subjected to LT and HH stress (9/5 °C day/night temperature, 95% humidity) as well as control (CK) conditions (25/18 °C day/night temperature, 80% humidity) for 24, 48, and 72 h. It was observed that the LT and HH stress caused severe damage to the morphometric traits of the plants compared to the control treatment. The concentrations of phytohormones IAA, ethylene, and GA were lower, while ABA and JA were higher during LT and HH stress at most time points. To gain insights into the molecular mechanisms underlying this stress response, RNA-sequencing was performed. The analysis revealed a total of 10,459 differentially expressed genes (DEGs) with annotated pathways. These pathways included plant hormone signal transduction, protein processing in the endoplasmic reticulum, MAPK signaling pathway, carbon fixation in photosynthetic organisms, and glycerolipid metabolism. Furthermore, 123 DEGs associated with hormone signaling pathways were identified, and their responses to LT and HH stress were thoroughly discussed. Overall, this study sheds light on the LT and HH tolerance mechanisms in cucumbers, particularly focusing on the genes involved in the LT and HH response and the signaling pathways of endogenous phytohormones.

Inhibitors against Two PDZ Domains of MDA-9 Suppressed Migration of Breast Cancer Cells.

Melanoma differentiation-associated gene 9 (MDA-9) is a small adaptor protein with tandem PDZ domains that promotes tumor progression and metastasis in various human cancers. However, it is difficult to develop drug-like small molecules with high affinity due to the narrow groove of the PDZ domains of MDA-9. Herein, we identified four novel hits targeting the PDZ1 and PDZ2 domains of MDA-9, namely PI1A, PI1B, PI2A, and PI2B, using a protein-observed nuclear magnetic resonance (NMR) fragment screening method. We also solved the crystal structure of the MDA-9 PDZ1 domain in complex with PI1B and characterized the binding poses of PDZ1-PI1A and PDZ2-PI2A, guided by transferred paramagnetic relaxation enhancement. The protein-ligand interaction modes were then cross-validated by the mutagenesis of the MDA-9 PDZ domains. Competitive fluorescence polarization experiments demonstrated that PI1A and PI2A blocked the binding of natural substrates to the PDZ1 and PDZ2 domains, respectively. Furthermore, these inhibitors exhibited low cellular toxicity, but suppressed the migration of MDA-MB-231 breast carcinoma cells, which recapitulated the phenotype of MDA-9 knockdown. Our work has paved the way for the development of potent inhibitors using structure-guided fragment ligation in the future.

Role of MCP-1 as an inflammatory biomarker in nephropathy.

The Monocyte chemoattractant protein-1 (MCP-1), also referred to as chemokine ligand 2 (CCL2), belongs to the extensive chemokine family and serves as a crucial mediator of innate immunity and tissue inflammation. It has a notable impact on inflammatory conditions affecting the kidneys. Upon binding to its receptor, MCP-1 can induce lymphocytes and NK cells' homing, migration, activation, differentiation, and development while promoting monocytes' and macrophages' infiltration, thereby facilitating kidney disease-related inflammation. As a biomarker for kidney disease, MCP-1 has made notable advancements in primary kidney diseases such as crescentic glomerulonephritis, chronic glomerulonephritis, primary glomerulopathy, idiopathic proteinuria glomerulopathy, acute kidney injury; secondary kidney diseases like diabetic nephropathy and lupus nephritis; hereditary kidney diseases including autosomal dominant polycystic kidney disease and sickle cell kidney disease. MCP-1 not only predicts the occurrence, progression, prognosis of the disease but is also closely associated with the severity and stage of nephropathy. When renal tissue is stimulated or experiences significant damage, the expression of MCP-1 increases, demonstrating a direct correlation with the severity of renal injury.

Melatonin Rescues Photosynthesis and Triggers Antioxidant Defense Response in Cucumis sativus Plants Challenged by Low Temperature and High Humidity.

Environmental factors such as low temperature (LT) and high humidity (HH) hinder plant growth and development in plastic tunnels and solar greenhouses in the cold season. In this study, we examined the effect of melatonin (MT) on shoot-based tolerance to LT and HH conditions in cucumber (Cucumis sativus) seedlings and explored its underlying mechanism. LT and HH stress inhibited growth and biomass accumulation, produced leaf chlorosis, led to oxidative stress, lowered chlorophyll and carotenoid contents, reduced photosynthetic and photosystem II (PSII) activities, and increased the level of intercellular carbon dioxide and the non-photochemical quenching of photosystem I (PSI) and PSII. However, foliar application of MT significantly improved the morphological indices and photosynthetic efficiency of cucumber seedlings, which entailed the elevation of electrolyte leakage, lipid peroxidation, and reactive oxygen species accumulation by boosting the antioxidant enzyme defense system under LT and HH conditions. Additionally, the measurement of nitrogen (N), magnesium (Mg), and iron (Fe) contents in roots and leaves showed that MT significantly augmented the nutrient uptake of cucumber seedlings exposed to LT and HH stresses. Furthermore, MT application increased the transcripts levels of genes encoding antioxidant enzymes under LT and HH conditions, whereas treatment with LT and HH suppressed these genes, suggesting that MT application increases the LT and HH tolerance of cucumber seedlings. Overall, our results suggest that MT application increases the tolerance of cucumber seedlings to LT and HH stress by enhancing the plant morphometric parameters, regulating PSI and PSII, and activating the antioxidant defense mechanism. Thus, the exogenous application of MT could be potentially employed as a strategy to improve the LT and HH tolerance of cucumber.

A Review of Traditional Chinese Medicine in Treating Renal Interstitial Fibrosis via Endoplasmic Reticulum Stress-Mediated Apoptosis.

Renal interstitial fibrosis (RIF) is the main pathological manifestation of end-stage renal disease. Recent studies have shown that endoplasmic reticulum (ER) stress is involved in the pathogenesis and development of RIF. Traditional Chinese medicine (TCM), as an effective treatment for kidney diseases, can improve kidney damage by affecting the apoptotic signaling pathway mediated by ER stress. This article reviews the apoptotic pathways mediated by ER stress, including the three major signaling pathways of unfolded protein response, the main functions of the transcription factor C/EBP homologous protein. We also present current research on TCM treatment of RIF, focusing on medicines that regulate ER stress. A new understanding of using TCM to treat kidney disease by regulating ER stress will promote clinical application of Chinese medicine and discovery of new drugs for the treatment of RIF.

Identification of a major-effect QTL associated with pre-harvest sprouting in cucumber (Cucumis sativus L.) using the QTL-seq method.

BACKGROUND: Cucumber (Cucumis sativus L.) is cultivated worldwide, and it is essential to produce enough high-quality seeds to meet demand. Pre-harvest sprouting (PHS) in cucumber is a critical problem and causes serious damage to seed production and quality. Nevertheless, the genetic basis and molecular mechanisms underlying cucumber PHS remain unclear. QTL-seq is an efficient approach for rapid quantitative trait loci (QTL) identification that simultaneously takes advantage of bulked-segregant analysis (BSA) and whole-genome resequencing. In the present research, QTL-seq analysis was performed to identify QTLs associated with PHS in cucumber using an F2 segregating population. RESULTS: Two QTLs that spanned 7.3 Mb on Chromosome 4 and 0.15 Mb on Chromosome 5 were identified by QTL-seq and named qPHS4.1 and qPHS5.1, respectively. Subsequently, SNP and InDel markers selected from the candidate regions were used to refine the intervals using the extended F2 populations grown in the 2016 and 2017 seasons. Finally, qPHS4.1 was narrowed to 0.53 Mb on chromosome 4 flanked by the markers SNP-16 and SNP-24 and was found to explain 19-22% of the phenotypic variation in cucumber PHS. These results reveal that qPHS4.1 is a major-effect QTL associated with PHS in cucumber. Based on gene annotations and qRT-PCR expression analyses, Csa4G622760 and Csa4G622800 were proposed as the candidate genes. CONCLUSIONS: These results provide novel insights into the genetic mechanism controlling PHS in cucumber and highlight the potential for marker-assisted selection of PHS resistance breeding.

Molecular Basis Underlying Leaf Variegation of a Moth Orchid Mutant (Phalaenopsis aphrodite subsp. formosana).

Leaf variegation is often the focus of plant breeding. Here, we studied a variegated mutant of Phalaenopsis aphrodite subsp. formosana, which is usually used as a parent of horticultural breeding, to understand its anatomic and genetic regulatory mechanisms in variegation. Chloroplasts with well-organized thylakoids and starch grains were found only in the mesophyll cells of green sectors but not of yellow sectors, confirming that the variegation belongs to the chlorophyll type. The two-dimensional electrophoresis and LC/MS/MS also reveal differential expressions of PsbP and PsbO between the green and yellow leaf sectors. Full-length cDNA sequencing revealed that mutant transcripts were caused by intron retention. When conditioning on the total RNA expression, we found that the functional transcript of PsbO and mutant transcript of PsbP are higher expressed in the yellow sector than in the green sector, suggesting that the post-transcriptional regulation of PsbO and PsbP differentiates the performance between green and yellow sectors. Because PsbP plays an important role in the stability of thylakoid folding, we suggest that the negative regulation of PsbP may inhibit thylakoid development in the yellow sectors. This causes chlorophyll deficiency in the yellow sectors and results in leaf variegation. We also provide evidence of the link of virus CymMV and the formation of variegation according to the differential expression of CymMV between green and yellow sectors.

Direct LAMP Assay without Prior DNA Purification for Sex Determination of Papaya.

Papaya (Carica papaya L.) is an economically important tropical fruit tree with hermaphrodite, male and female sex types. Hermaphroditic plants are the major type used for papaya production because their fruits have more commercial advantages than those of female plants. Sex determination of the seedlings, or during the early growth stages, is very important for the papaya seedling industry. Thus far, the only method for determining the sex type of a papaya at the seedling stage has been DNA analysis. In this study, a molecular technique-based on DNA analysis-was developed for detecting male-hermaphrodite-specific markers to examine the papaya's sex type. This method is based on the loop-mediated isothermal amplification (LAMP) and does not require prior DNA purification. The results show that the method is an easy, efficient, and inexpensive way to determine a papaya's sex. This is the first report on the LAMP assay, using intact plant materials-without DNA purification-as samples for the analysis of sex determination of papaya. We found that using high-efficiency DNA polymerase was essential for successful DNA amplification, using trace intact plant material as a template DNA source.

Comparative transcriptome analysis of Gastrodia elata (Orchidaceae) in response to fungus symbiosis to identify gastrodin biosynthesis-related genes.

BACKGROUND: Gastrodia elata Blume (Orchidaceae) is an important Chinese medicine with several functional components. In the life cycle of G. elata, the orchid develops a symbiotic relationship with two compatible mycorrhizal fungi Mycena spp. and Armillaria mellea during seed germination to form vegetative propagation corm and vegetative growth to develop tubers, respectively. Gastrodin (p-hydroxymethylphenol-beta-D-glucoside) is the most important functional component in G. elata, and gastrodin significantly increases from vegetative propagation corms to tubers. To address the gene regulation mechanism in gastrodin biosynthesis in G. elata, a comparative analysis of de novo transcriptome sequencing among the vegetative propagation corms and tubers of G. elata and A. mellea was conducted using deep sequencing. RESULTS: Transcriptome comparison between the vegetative propagation corms and juvenile tubers of G. elata revealed 703 differentially expressed unigenes, of which 298 and 405 unigenes were, respectively up-regulated (fold-change ≥ 2, q-value < 0.05, the trimmed mean of M-values (TMM)-normalized fragments per kilobase of transcript per Million mapped reads (FPKM) > 10) and down-regulated (fold-change ≤ 0.5, q-value <0.05, TMM-normalized FPKM > 10) in juvenile tubers. After Gene Ontology (GO) annotation and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, 112 up-regulated unigenes with KEGG Ortholog identifiers (KOids) or enzyme commission (EC) numbers were assigned to 159 isogroups involved in seventy-eight different pathways, and 132 down-regulated unigenes with KOids or EC numbers were assigned to 168 isogroups, involved in eighty different pathways. The analysis of the isogroup genes from all pathways revealed that the two unigenes TRINITY_DN54282_c0_g1 (putative monooxygenases) and TRINITY_DN50323_c0_g1 (putative glycosyltransferases) might participate in hydroxylation and glucosylation in the gastrodin biosynthetic pathway. CONCLUSIONS: The gene expression of the two unique unigenes encoding monooxygenase and glycosyltransferase significantly increases from vegetative propagation corms to tubers, and the molecular basis of gastrodin biosynthesis in the tubers of G. elata is proposed.

The effect of biocontrol bacteria on rhizosphere bacterial communities analyzed by plating and PCR-DGGE.

Bacillus subtilis B579, which was isolated from rhizosphere of cucumber, exhibited an excellent biocontrol activity on soil-born pathogens under greenhouse conditions. It could colonize in rhizosphere of cucumber with large number of populations after inoculated in plant growth season. To reveal the effect of high level colonization of B. subtilis B579 on rhizobacteria community structure, cultivation-based analysis coupled with denaturing gradient gel electrophoresis (DGGE) analysis were used to profile the changes of rhizobacteria community structure sampling at 1 week interval. Cultivation-based and DGGE fingerprinting analysis showed significant plant-dependent and seasonal shifts in rhizobacteria populations. Only minimal and transient effects were observed at 4-9 weeks after sowing in samples of B579 treatment, without the pathogen inoculation and showed the best plant growth potential. Sequencing of dominant bands excised from the gel revealed that Streptomyces sp. was the dominate species in soils before and after sowing. Burkholderia sp. was the dominate species in bulk soil, while Bacillus sp. was dominated in rhizosphere within the growth season. Arthrobacter ramosus and Nocardioides sp. were identified as the specific species in samples treated by B579 at the maturity and flowering stages of cucumber.