Upregulation of CALD1 predicted a poor prognosis for platinum-treated ovarian cancer and revealed it as a potential therapeutic resistance target.

BACKGROUND: Ovarian cancer (OC) has the worst prognosis among gynecological malignancies, most of which are found to be in advanced stage. Cell reduction surgery based on platinum-based chemotherapy is the current standard of treatment for OC, but patients are prone to relapse and develop drug resistance. The objective of this study was to identify a specific molecular target responsible for platinum chemotherapy resistance in OC. RESULTS: We screened the protein-coding gene Caldesmon (CALD1), expressed in cisplatin-resistant OC cells in vitro. The prognostic value of CALD1 was evaluated using survival curve analysis in OC patients treated with platinum therapy. The diagnostic value of CALD1 was verified by drawing a Receiver Operating Characteristic (ROC) curve using clinical samples from OC patients. This study analyzed data from various databases including Gene Expression Omnibus (GEO), Human Protein Atlas (HPA), The Cancer Cell Line Encyclopedia (CCLE), The Cancer Genome Atlas (TCGA), GEPIA 2, UALCAN, Kaplan-Meier (KM) plotter, LinkedOmics database, and String. Different expression genes (DEGs) between cisplatin-sensitive and cisplatin-resistant cells were acquired respectively from 5 different datasets of GEO. CALD1 was selected as a common gene from 5 groups DEGs. Online data analysis of HPA and CCLE showed that CALD1 was highly expressed in both normal ovarian tissue and OC. In TCGA database, high expression of CALD1 was associated with disease stage and venous invasion in OC. Patients with high CALD1 expression levels had a worse prognosis under platinum drug intervention, according to Kaplan-Meier (KM) plotter analysis. Analysis of clinical sample data from GEO showed that CALD1 had superior diagnostic value in distinguishing patients with platinum "resistant" and platinum "sensitive" (AUC = 0.816), as well as patients with worse progression-free survival (AUC = 0.741), and those with primary and omental metastases (AUC = 0.811) in ovarian tumor. At last, CYR61 was identified as a potential predictive molecule that may play an important role alongside CALD1 in the development of platinum resistance in OC. CONCLUSIONS: CALD1, as a member of cytoskeletal protein, was associated with poor prognosis of platinum resistance in OC, and could be used as a target protein for mechanism study of platinum resistance in OC.

Brassinosteroids is involved in methane-induced adventitious root formation via inducing cell wall relaxation in marigold.

BACKGROUND: Methane (CH4) and brassinosteroids (BRs) are important signaling molecules involved in a variety of biological processes in plants. RESULTS: Here, marigold (Tagetes erecta L. 'Marvel') was used to investigate the role and relationship between CH4 and BRs during adventitious root (AR) formation. The results showed a dose-dependent effect of CH4 and BRs on rooting, with the greatest biological effects of methane-rich water (MRW, CH4 donor) and 2,4-epibrassinolide (EBL) at 20% and 1 µmol L- 1, respectively. The positive effect of MRW on AR formation was blocked by brassinoazole (Brz, a synthetic inhibitor of EBL), indicating that BRs might be involved in MRW-regulated AR formation. MRW promoted EBL accumulation during rooting by up-regulating the content of campestanol (CN), cathasterone (CT), and castasterone (CS) and the activity of Steroid 5α-reductase (DET2), 22α-hydroxylase (DWF4), and BR-6-oxidase (BR6ox), indicating that CH4 could induce endogenous brassinolide (BR) production during rooting. Further results showed that MRW and EBL significantly down-regulated the content of cellulose, hemicellulose and lignin during rooting and significantly up-regulated the hydrolase activity, i.e. cmcase, xylanase and laccase. In addition, MRW and EBL also significantly promoted the activity of two major cell wall relaxing factors, xyloglucan endotransglucosylase/hydrolase (XTH) and peroxidase, which in turn promoted AR formation. While, Brz inhibited the role of MRW on these substances. CONCLUSIONS: BR might be involved in CH4-promoted AR formation by increasing cell wall relaxation.

A spatiotemporal steroidogenic regulatory network in human fetal adrenal glands and gonads.

Human fetal adrenal glands produce substantial amounts of dehydroepiandrosterone (DHEA), which is one of the most important precursors of sex hormones. However, the underlying biological mechanism remains largely unknown. Herein, we sequenced human fetal adrenal glands and gonads from 7 to 14 gestational weeks (GW) via 10× Genomics single-cell transcriptome techniques, reconstructed their location information by spatial transcriptomics. Relative to gonads, adrenal glands begin to synthesize steroids early. The coordination among steroidogenic cells and multiple non-steroidogenic cells promotes adrenal cortex construction and steroid synthesis. Notably, during the window of sexual differentiation (8-12 GW), key enzyme gene expression shifts to accelerate DHEA synthesis in males and cortisol synthesis in females. Our research highlights the robustness of the action of fetal adrenal glands on gonads to modify the process of sexual differentiation.

Involvement of Calcium and Calmodulin in NO-Alleviated Salt Stress in Tomato Seedlings.

Salt stress is an adverse impact on the growth and development of plants, leading to yield losses in crops. It has been suggested that nitric oxide (NO) and calcium ion (Ca2+) act as critical signals in regulating plant growth. However, their crosstalk remains unclear under stress condition. In this study, we demonstrate that NO and Ca2+ play positive roles in the growth of tomato (Lycopersicum esculentum) seedlings under salt stress. Our data show that Ca2+ channel inhibitor lanthanum chloride (LaCl3), Ca2+ chelator ethylene glycol-bis (2-aminoethylether)-N,N,N,N-tetraacetic acid (EGTA), or calmodulin (CaM) antagonist N-(6-aminohexyl)-5-chloro-1-naphthalenesulfona-mide hydrochloride (W-7) significantly reversed the effect of NO-promoted the growth of tomato seedlings under salt stress. We further show that NO and Ca2+ significantly decreased reactive oxygen accumulation, increased proline content, and increased the activity of antioxidant enzymes, as well as increased expression of antioxidant enzymes related genes. However, LaCl3, EGTA, and W-7 prevented the positive roles of NO. In addition, the activity of downstream target enzymes related to Ca2+/CaM was increased by NO under salt stress, while LaCl3, EGTA, and W-7 reversed this enhancement. Taken together, these results demonstrate that Ca2+/CaM might be involved in NO-alleviate salt stress.

Abscisic Acid Induces Adventitious Rooting in Cucumber (Cucumis sativus L.) by Enhancing Sugar Synthesis.

Abscisic acid (ABA) affects many important plant processes, such as seed germination, root elongation and stomatal movement. However, little information is available about the relationship between ABA and sugar synthesis during adventitious root formation. The aim of this study was to evaluate the effect of ABA on adventitious root formation in cucumber and whether the effect of this plant hormone on sugar synthesis could be included as a causative factor for adventitious root development. We determined the contents of glucose, sucrose, starch, total sugar and sugar-related enzymes, including sucrose synthase (SS), sucrose phosphate synthase (SPS), hexokinase (HK) and pyruvate kinase (PK) activities in ABA treatment. We also quantified the relative expression of sucrose or glucose synthesis genes during this process. Increasing ABA concentrations significantly improved adventitious root formation, with the most considerable effect at 0.05 µM. Compared to the control, ABA treatment showed higher glucose, sucrose, starch and total sugar contents. Moreover, ABA treatment increased glucose-6-phosphate (G6P), fructose-6-phosphate (F6P) and glucose-1-phosphate (G1P) contents in cucumber explants during adventitious root development, which was followed by an increase of activities of sucrose-related enzymes SS and SPS, glucose-related enzymes HK and PK. ABA, meanwhile, upregulated the expression levels of sucrose or glucose synthesis-related genes, including CsSuSy1, CsSuSy6, CsHK1 and CsHK3. These results suggest that ABA may promote adventitious root development by increasing the contents of glucose, sucrose, starch, total sugar, G6P, F6P and G1P, the activities of SS, SPS, HK, SPS and the expression levels of CsSuSy1, CsSuSy6, CsHK1 and CsHK3 genes. These findings provide evidence for the physiological role of ABA during adventitious root formation and provide a new understanding of the possible relationship between ABA and sugar synthesis during adventitious rooting in plants.

Nitric Oxide Enhanced Salt Stress Tolerance in Tomato Seedlings, Involving Phytohormone Equilibrium and Photosynthesis.

Nitric oxide (NO), as a ubiquitous gas signaling molecule, modulates various physiological and biochemical processes and stress responses in plants. In our study, the NO donor nitrosoglutathione (GSNO) significantly promoted tomato seedling growth under NaCl stress, whereas NO scavenger 2-(4-carboxyphenyl)-4, 4, 5, 5-tetramethylimidazoline-1-oxyl-3-oxide potassium (cPTIO) treatment reversed the positive effect of NO, indicating that NO plays an essential role in enhancing salt stress resistance. To explore the mechanism of NO-alleviated salt stress, the transcriptome of tomato leaves was analyzed. A total of 739 differentially expressed genes (DEGs) were identified and classified into different metabolic pathways, especially photosynthesis, plant hormone signal transduction, and carbon metabolism. Of these, approximately 16 and 9 DEGs involved in plant signal transduction and photosynthesis, respectively, were further studied. We found that GSNO increased the endogenous indoleacetic acid (IAA) and salicylic acid (SA) levels but decreased abscisic acid (ABA) and ethylene (ETH) levels under salt stress conditions. Additionally, GSNO induced increases in photosynthesis pigment content and chlorophyll fluorescence parameters under NaCl stress, thereby enhancing the photosynthetic capacity of tomato seedlings. Moreover, the effects of NO mentioned above were reversed by cPTIO. Together, the results of this study revealed that NO regulates the expression of genes related to phytohormone signal transduction and photosynthesis antenna proteins and, therefore, regulates endogenous hormonal equilibrium and enhances photosynthetic capacity, alleviating salt toxicity in tomato seedlings.

An optimized method to obtain high-quality RNA from different tissues in Lilium davidii var. unicolor.

The high quality, yield and purity total RNA samples are essential for molecular experiments. However, harvesting high quality RNA in Lilium davidii var. unicolor is a great challenge due to its polysaccharides, polyphenols and other secondary metabolites. In this study, different RNA extraction methods, namely TRIzol method, the modified TRIzol method, Kit method and cetyltrimethylammonium bromide (CTAB) method were employed to obtain total RNA from different tissues in L. davidii var. unicolor. A Nano drop spectrophotometer and 1% agarose gel electrophoresis were used to detect the RNA quality and integrity. Compared with TRIzol, Kit and CTAB methods, the modified TRIzol method obtained higher RNA concentrations from different tissues and the A260/A280 ratios of RNA samples were ranged from 1.97 to 2.27. Thus, the modified TRIzol method was shown to be the most effective RNA extraction protocol in acquiring RNA with high concentrations. Furthermore, the RNA samples isolated by the modified TRIzol and Kit methods were intact, whereas different degrees of degradation happened within RNA samples isolated by the TRIzol and CTAB methods. In addition, the modified TRIzol method could also isolate high-quality RNA from other edible lily bulbs. Taken together, the modified TRIzol method is an efficient method for total RNA isolation from L. davidii var. unicolor.

Hydrogen-rich water promotes the formation of bulblets in Lilium davidii var. unicolor through regulating sucrose and starch metabolism.

MAIN CONCLUSION: HRW increased the content of starch and sucrose via regulating a series of sucrose and starch synthesis genes, which induced the formation of bulblets and adventitious roots of Lilium davidii var. unicolor. Hydrogen gas (H2), as a signaling molecule, has been reported to be involved in plant growth and development. Here, the effect of hydrogen-rich water (HRW) on the formation of bulblets and adventitious roots in the scale cuttings of Lilium davidii var. unicolor and its mechanisms at the molecular levels were investigated. The results revealed that compared with distilled water treatment (Con), the number of bulblets and adventitious roots were significantly promoted by different concentrations of HRW treatment. Treatment with 100% HRW obtained the most positive effects. RNA sequencing (RNA-seq) analysis found that compared with Con, a total of 1702 differentially expressed genes (DEGs, upregulated 552 DEGs, downregulated 1150 DEGs) were obtained under HRW treatment. The sucrose and starch metabolism, cysteine and methionine metabolism and phenylalanine metabolism were significantly enriched in the analysis of the Kyoto encyclopedia of genes and genomes (KEGG). In addition, the genes involved in carbohydrate metabolism were significantly upregulated or downregulated (upregulated 22 DEGs, downregulated 15 DEGs), indicating that starch and sucrose metabolism held a central position. The expressions of 12 DEGs were identified as coding for key enzymes in metabolism of carbohydrates was validated by qPCR during bulblet formation progress. RNA-seq analysis and expression profiles indicated that the unigene levels such as glgc, Susy, otsA and glgP, BMY and TPS were well correlated with sucrose and starch metabolism during HRW-induced bulblet formation. The change of key enzyme content in starch and sucrose metabolism pathway was explored during bulblet formation in Lilium under HRW treatment. Meanwhile, compared with Con, 100% HRW treatment increased the levels of sucrose and starch, and decreased the trehalose content, which were agreed with the expression pattern of DEGs related to the biosynthesis pathway of sucrose, starch and trehalose. Therefore, this study suggested that HRW could promote the accumulation of sucrose and starch contents in mother scales, and decreased the trehalose content, this might provide more energy for bulblet formation.

The Role of Osteocalcin in Placental Function in Gestational Diabetes Mellitus.

Evidence for the role of osteocalcin in glucose metabolism is increasing. The aim of this study was to examine the associations between osteocalcin and gestational diabetes mellitus. Thirteen discovery study subjects and 76 reduplication study subjects were recruited from the Maternal and Child Health Hospital Guangxi Zhuang Autonomous Region from May 2018 to August 2018. Total osteocalcin and biochemical indices of maternal serum and umbilical vein serum were analyzed. Placental tissue samples were used for transcriptome sequencing. For the discovery study subjects, the total osteocalcin concentration in umbilical vein serum was significantly higher than that in maternal serum and umbilical artery serum (55.32 ng/mL ± 17.37 vs. 12.06 ng/mL ± 5.42 [P < 0.001] vs. 38.31 ng/mL ± 11.52 [P < 0.01]), suggesting that trophoblasts may synthesize osteocalcin. In a reduplication subject study, the gestational diabetes mellitus group had lower umbilical vein serum total osteocalcin (51.46 ng/mL ± 24.29 vs. 67.00 ng/mL ± 25.33, P = 0.008), lower adiponectin (1099.72 µg/L ± 102.65 vs. 1235.85 µg/L ± 94.63, P < 0.001). Spearman's correlation analysis showed that umbilical vein serum total osteocalcin levels were closely correlated with leptin (r = -0.456, P = 0.007). A coexpression model of the placental RNA sequence was constructed. Two modules were correlated with osteocalcin, and the Gene ontology pathways of these modules were rich in glucose and lipid metabolism. In conclusion, the placenta may synthesize osteocalcin by itself, and a lower osteocalcin level in umbilical vein serum is associated with gestational diabetes mellitus.

Methane-rich water induces bulblet formation of scale cuttings in Lilium davidii var. unicolor by regulating the signal transduction of phytohormones and their levels.

Previous studies have shown that methane (CH4 ) has promoting roles in the adventitious root (AR) and lateral root (LR) formation in plants. However, whether CH4 could trigger the bulblet formation in scale cutting of Lilium davidii var. unicolor has not been elucidated. To gain insight into the effect of CH4 on the bulblet formation, different concentrations (1, 10, 50, and 100%) of methane-rich water (MRW) and distilled water were applied to treat the scale cuttings of Lilium. We observed that treatment with 100% MRW obviously induced the bulblet formation in scale cuttings. To explore the mechanism of CH4 -induced bulblet formation, the transcriptome of scales was analyzed. A total of 2078 differentially expressed genes (DEGs) were identified. The DEGs were classified into different metabolic pathways, especially phenylpropanoid biosynthesis, starch and sucrose metabolism, and plant signal transduction. Of these, approximately 38 candidate DEGs involved in the plant signal transduction were further studied. In addition, the expression of AP2-ERF/ERF, WRKY, GRAS, ARF, and NAC transcription factors (TFs) was changed by MRW treatment, suggesting their potential involvement in bulblet formation. As for hormones, exogenous IAA, GA, and ABA could induce the bulblet formation. Additional experiments suggested that MRW could increase the endogenous IAA, GA, and JA levels, but decrease the levels of ABA during bulblet formation, which showed that higher IAA, GA, JA levels and lower ABA content might facilitate bulblet formation. In addition, the levels of endogenous hormones were consistent with the expression level of genes involved in phytohormone signal transduction. Overall, this study has revealed that CH4 might improve the bulblet formation of cutting scales in Lilium by regulating the expression of genes related to phytohormone signal transduction and TFs, as well as by changing the endogenous hormone levels.

The Endogenous Alterations of the Gut Microbiota and Feces Metabolites Alleviate Oxidative Damage in the Brain of LanCL1 Knockout Mice.

Altered composition of the gut microbiota has been observed in many neurodegenerative diseases. LanCL1 has been proven to protect neurons and reduce oxidative stress. The present study was designed to investigate alterations of the gut microbiota in LanCL1 knockout mice and to study the interactions between gut bacteria and the brain. Wild-type and LanCL1 knockout mice on a normal chow diet were evaluated at 4 and 8-9 weeks of age. 16s rRNA sequence and untargeted metabolomics analyses were performed to investigate changes in the gut microbiota and feces metabolites. Real-time polymerase chain reaction analysis, AB-PAS staining, and a TUNEL assay were performed to detect alterations in the gut and brain of knockout mice. The serum cytokines of 9-week-old knockout mice, which were detected by a multiplex cytokine assay, were significantly increased. In the central nervous system, there was no increase of antioxidant defense genes even though there was only low activity of glutathione S-transferase in the brain of 8-week-old knockout mice. Interestingly, the gut tight junctions, zonula occludens-1 and occludin, also displayed a downregulated expression level in 8-week-old knockout mice. On the contrary, the production of mucus increased in 8-week-old knockout mice. Moreover, the compositions of the gut microbiota and feces metabolites markedly changed in 8-week-old knockout mice but not in 4-week-old mice. Linear discriminant analysis and t-tests identified Akkermansia as a specific abundant bacteria in knockout mice. Quite a few feces metabolites that have protective effects on the brain were reduced in 8-week-old knockout mice. However, N-acetylsphingosine was the most significant downregulated feces metabolite, which may cause the postponement of neuronal apoptosis. To further investigate the effect of the gut microbiota, antibiotics treatment was given to both types of mice from 5 to 11 weeks of age. After treatment, a significant increase of oxidative damage in the brain of knockout mice was observed, which may have been alleviated by the gut microbiota before. In conclusion, alterations of the gut microbiota and feces metabolites alleviated oxidative damage to the brain of LanCL1 knockout mice, revealing that an endogenous feedback loop mechanism of the microbiota-gut-brain axis maintains systemic homeostasis.

rs2274911 polymorphism in GPRC6A associated with serum E2 and PSA in a Southern Chinese male population.

BACKGROUND: rs2274911 (Pro91Ser, G > A) is a missense mutation located on the second exon of the GPRC6A gene. Increasing evidence revealed a significant association between the A allele of rs2274911 and male diseases, such as oligospermia, cryptorchidism, and prostate tumor. However, the function of rs2274911 in healthy males is unclear. SUBJECTS AND METHODS: A total of 1742 healthy men were selected from the Fangchenggang Area Male Health and Examination Survey (FAMHES). The association between rs2274911 and phenotype was evaluated. The cell characteristics of rs2274911 mutation (mu), wild-type GPRC6A (WT), and RFP control in human embryonic kidney (293T) and human prostate cancer (PC3) cells were analyzed. RNA sequencing was performed on PC3 cells. RESULTS: E2 and PSA serum levels increased with the accumulation of the A allele (E2: G vs. A, -0.029 [-0.050, -0.008], P < 0.01, P trend = 0.027; PSA: G vs. A, -0.040 [-0.079, 0.000], P < 0.05, P trend = 0.048). rs2274911 enhanced the proliferation and invasion ability of PC3 or 293T cells and activated the ERK pathway. The genes were identified as rs2274911 mu-affected genes through RNA sequential analysis of rs2274911 mu, GPRC6A WT, and RFP control of PC3 cells. Most of these genes were related to cancer development processes, cAMP, and the ERK cell signaling pathway. CONCLUSION: This project represents that rs2274911 is associated with E2 and PSA serum levels in Southern Chinese men. Rs2274991 mutation promotes 293T and PC3 cell proliferation in vitro. These results suggest that rs2274911 is a functional variant of GPRC6A.

FTO Facilitates Lung Adenocarcinoma Cell Progression by Activating Cell Migration Through mRNA Demethylation.

BACKGROUND: The fat mass and obesity-associated protein (FTO) was identified as a critical demethylase involved in regulating cellular mRNA stability by removing N6-methyladenosine (m6A) residues from mRNA. Emerging evidence has revealed that FTO is deeply implicated in lung cancer. However, knowledge of the function of FTO in lung adenocarcinoma (LUAC) is limited. METHODS: FTO and FTO R96Q (R96Q), an FTO missense mutant lacking demethylase activity, were ectopically overexpressed, and FTO was knocked down via siRNA in A549 and H1299 cells. The relationships between FTO with cell characteristics and mRNA m6A levels were explored. Furthermore, RNA sequencing was performed on A549 cells. RESULTS: FTO overexpression enhanced the proliferation, migration, and invasion ability of A549 and H1299 cells, decreased mRNA m6A levels. Interestingly, overexpression of R96Q, blunted the effects of FTO overexpression on cell proliferation and invasion. Through RNA sequencing analysis of A549 cells overexpressing FTO or R96Q and control A594 cells, 45 genes were identified as affected by m6A mRNA demethylation. Most of these genes were related to lung cancer, such as laminin γ2, thrombospondin 1, nerve growth factor inducible, integrin alpha11, and proprotein convertase subtilisin/kexin type 9. Gene ontology and Kyoto Encyclopedia of Genes and Genomes analyses suggested that these genes are fundamental to cancer development processes, such as cell migration and extracellular matrix organization. CONCLUSION: Our research shows that FTO facilitates LUAC cell progression by activating cell migration through m6A demethylation; however, further research on the mechanism underlying FTO activity in LUAC is necessary.

Unchanged Pulmonary Function and Increased Prevalence of Subjective Respiratory Symptoms in Non-smoking Workers with Low Cadmium Body Burden.

The study investigated the effects of low cadmium (Cd) burden on the respiratory symptoms and pulmonary function in occupational workers. The study population consisted of 98 nickel-cadmium battery workers. Levels of urine cadmium, urinary creatinine (Cr), and data on the adverse respiratory symptoms and pulmonary function of the participants were measured and collected respectively. The urinary cadmium level in cadmium-exposed workers (n = 53) was within the normal range but greater than 2.4 times than those of the control group (n = 45). Compared with the control group, the cadmium-exposed workers had higher prevalence of all subjective respiratory symptoms studied in the study. The prevalence of cough (30%), phlegm (23%), and upper respiratory tract infection (URTI) (79%) in the exposure group was significantly (P < 0.05) higher than the prevalence of cough (11%), phlegm (7%), and URTI (31%) in the control group respectively. There was no significant difference between the Cd-exposed workers and control group in pulmonary function test. Our findings suggested that adverse subjective respiratory symptoms were increased and pulmonary function were unchanged in low Cd burden workers. Unchanged pulmonary function may be related to age, exposure duration, and distribution of Cd in tissue. These workers need to continue being monitored in longitudinal studies.

[Toxic effects of mixture of volatile organic compounds on mice testis related enzymes and hormones].

OBJECTIVE: To evaluate the toxic effects of mixture of volatile organic compounds (VOCs) on Mice Testis related enzymes and hormones. METHODS: After determining the median lethal dose (LD50) of VOCs using the acute toxicity test, 40 male clean inbred Kunming mice were assigned to 1/8 LD50 VOCs exposure group, 1/4 LD50 VOCs exposure group, and 1/2 LD50 VOCs exposure group, as well as positive control group with cyclophosphamide (60 mg/kg) and negative control group with tea oil, with 8 mice in each group. The mice were intraperitoneally injected with respective agents for 5 days. The levels of testis testosterone, estradiol, follicle stimulating hormone, and luteinizing hormone were determined by ELISA. Meanwhile, the activity of testicular marked enzymes such as lactate dehydrogenase, gamma-glutamyl transpeptidase, acid phosphatase, and glucose-6-phosphate dehydrogenase were examined. RESULTS: Compared with the negative control group, the 1/8 LD50 exposure group had a significantly increased testis coefficient (P<0.05). Both the activity of testicular marked enzymes and the levels of testicular sex hormones in all exposure groups showed significant downward trends with increasing VOC doses compared with those in the negative control group (P<0.05). CONCLUSION: VOCs have obvious toxicity to mouse testis by changing the levels of testicular sex hormones and the activity of testicular marked enzymes.