The nod-like receptor family pyrin domain containing 3 (NLRP3) inflammasome plays a crucial role in the prognosis of subarachnoid hemorrhage (SAH). WNK1 kinase negatively regulates NLRP3 in various inflammatory conditions, but its role in early brain injury (EBI) after SAH remains unclear. In this study, we used an in vivo SAH model in rats/mice and AAV-WNK1 intraventricular injection to investigate its neuroprotective mechanisms. WNK1 expression was significantly reduced in SAH patient blood and SAH model brain tissue, correlating negatively with microglial activation. AAV-WNK1 alleviated brain edema, neuronal necrosis, behavioral deficits, and inflammation by inhibiting NLRP3 inflammasome activation. In hemin-stimulated BV-2 cells, WNK1 overexpression reduced NLRP3 activation and inflammatory cytokines. Chloride counteracted WNK1's inhibitory effects, and WNK1 suppressed P2X7R-induced NLRP3 activation. Mechanistically, WNK1 functioned via the OXSR1/STK39 pathway. These findings highlight WNK1 as a key regulator of intracellular chloride balance and neuroinflammation, presenting a potential therapeutic target for SAH treatment.
Animals ; NLR Family, Pyrin Domain-Containing 3 Protein/metabolism* ; Subarachnoid Hemorrhage/complications* ; Inflammasomes/metabolism* ; Rats ; Mice ; Neuroinflammatory Diseases/metabolism* ; WNK Lysine-Deficient Protein Kinase 1/genetics* ; Male ; Humans ; Chlorides/metabolism* ; Mice, Inbred C57BL ; Rats, Sprague-Dawley ; Brain Injuries/metabolism* ; Microglia/metabolism* ; Protein Serine-Threonine Kinases

Objective To analyze the factors influencing the deterioration of nutritional status after radiotherapy/radiochemotherapy for esophageal cancer,so as to provide reference for nutritional management during antitumor therapy.Methods A total of 106 patients with esophageal cancer who received radiotherapy or radiochemotherapy at Anqing First People's Hospital of Anhui Medical University from Dec.2017 to Dec.2023 were enrolled.Patients'gender,age,surgical history,timing of radiotherapy intervention,synchronous chemoradiotherapy,radiotherapy dose,clinical stage,initial nutritional status,and performance status score were collected.The patient generated subjective global assessment scale(PG-SGA)scores were monitored before and after antitumor treatment.According to the nutritional status at the beginning of enrollment and at the end of radiotherapy,the patients were assigned to deterioration group or non-deterioration(stable or improved)group.The clinical characteristics of the 2 groups were compared.The factors influencing the deterioration of nutritional status were screened by logistic regression analysis.The correlation between nutritional status deterioration and adverse reactions(radiation esophagitis,pulmonary infection,neutropenia,thrombocytopenia,and elevated aminotransferase)was analyzed by Spearman correlation analysis.Results There were no significant differences in gender,radiotherapy dose,initial nutritional status,or performance status score between the 2 groups for the deterioration of nutritional status after radiotherapy(all P＞0.05).The proportions of patients with previous surgical history of esophageal cancer,synchronous chemoradiotherapy,initiation of radiotherapy at less than 90%of target calorie requirement,and clinical stage Ⅳ were significantly higher in the deterioration group than those in the non-deterioration group(all P＜0.05).Logistic regression analysis showed that clinical stage Ⅳ(odds ratio[OR]=4.684,95%confidence interval[CI]1.252-17.519,P=0.022)and previous surgical history of esophageal cancer(OR=7.338,95%CI 1.878-28.666,P=0.004)were the independent adverse risk factors for the deterioration of nutritional status after radiotherapy/radiochemotherapy.The timing of radiotherapy intervention was also an independent risk factor for the deterioration of nutritional status,and taking the tolerance of 70%-90%target energy as the reference level,starting radiotherapy when the tolerance of 90%-100%target energy had the optimal protection of nutritional status(OR=0.166,95%CI 0.050-0.551,P=0.003).Spearman correlation analysis showed that the deterioration of nutritional status was positively correlated with elevated transaminases after radiotherapy(rs=0.283,P=0.003),while it was not correlated with the other adverse reactions(all P＞0.05).Conclusion Under the standard nutritional intervention model,patients with previous surgery and recurrent metastatic esophageal cancer who receive radiotherapy/chemoradiotherapy are still at risk of nutritional status deterioration.Tolerance to 90%-100%target energy requirement may be a more appropriate timing for radiotherapy intervention.When the nutritional status deteriorates during treatment,it is necessary to be alert to the elevated transaminases.

Objective To investigate the relationship of miRNA gene polymorphisms with blood pressure(BP)responses to the sodium and potassium diet intervention.Methods In 2004,we recruited 514 participants from 124 families in seven villages of Baoji,Shaanxi Province,China.All subjects were given a three-day normal diet,followed by a seven-day low-salt diet,a seven-day high-salt diet,and finally a seven-day high-salt and potassium supplementation.A total of 19 miRNA single nucleotide polymorphisms(SNPs)were selected for analysis.Results Throughout the sodium-potassium dietary intervention,the BP of the subjects fluctuated across all phases,showing a decrease during the low-salt period and an increase during the high-salt period,followed by a reduction in BP subsequent to potassium supplementation during the high-salt diet.MiR-210-3p SNP rs 12364149 was significantly associated with systolic BP(SBP),diastolic BP(DBP)and mean arterial pressure(MAP)responses to low-salt diet.MiR-4638-3p SNP rs6601178 was significantly associated with SBP while miR-26b-3p SNP rs115254818 was significantly associated with MAP responses to low-salt intervention.In addition,miR-26b-3p SNP rs115254818 was significantly correlated with SBP,DBP and MAP responses to high-salt intervention.MiR-1307-5p SNPs rs1 1191676 and rs2292807 were associated with SBP and MAP responses to high-salt diet.MiR-4638-3p SNP rs6601178,miR-210-3p SNP rs12364149,miR-382-5p SNP rs4906032 and rs4143957 were significantly associated with SBP response to high-salt diet.In addition,miR-26b-3p SNP rs115254818 was significantly associated with SBP,DBP and MAP responses to potassium supplementation.MiR-1307-5p SNPs rs11191676,rs2292807,and miR-19a-3p SNP rs4284505 were significantly associated with SBP responses to high-salt and potassium supplementation.Conclusion miRNA gene polymorphisms are associated with BP response to sodium and potassium,suggesting that miRNA genes may be involved in the pathophysiological process of salt sensitivity and potassium sensitivity.

Objective:To evaluate the relationship between hippocampal receptor-interacting protein kinase-1 (RIPK1) and nucleotide-binding oligomerization domain-like receptor containing pyrin domain 3 (NLRP3) inflammasomes in postoperative neurocognitive dysfunction of aged rats with chronic knee arthritis pain.Methods:Sixty-four healthy male Sprague-Dawley rats, aged 18 months, weighing 500-550 g, were divided into 4 groups ( n=16 each) using a random number table method: chronic knee arthritis pain group (group P), chronic knee arthritis pain+ operation group (group PS), RIPK1 inhibitor necrostatin-1+ chronic knee arthritis pain+ operation group (group NPS), and DMSO+ chronic knee arthritis pain+ operation group (group DPS). The knee arthritis model was prepared by intra-articular injection of monosodium iodoacetate (MIA) 1 mg into the left knee joint, and 12 weeks later exploratory laparotomy was performed under sevoflurane anesthesia. Necrostatin-1 6.25 mg/kg and the equal volume of DMSO were intraperitoneally injected at 1 h before operation in NPS group and DPS group, respectively. Thermal pain threshold was measured at 1 week before MIA injection and 6 and 12 weeks after MIA injection. Morris water maze test was used to evaluate the cognitive function at 7 days after surgery. Hippocampal tissues were obtained for microscopic examination of the pathological changes (after HE staining) and for determination of the expression of RIPK1, phosphorylated RIPK1 (p-RIPK1), NLRP3, activated cysteine-aspartic protease caspase-1 (cl-caspase-1), apoptosis-associated speck-like protein containing a CARD (ASC) (by Western blot) and contents of interleukin-1beta (IL-1β) and IL-18 (by enzyme-linked immunosorbent assay). Results:Thermal pain threshold was significantly decreased at 6 and 12 weeks after MIA injection as compared with that before injection ( P<0.05), and there was no significant difference in thermal pain threshold among the four groups ( P>0.05). Compared with P group, the escape latency was significantly prolonged, the time of staying at the original platform quadrant was shortened, the number of crossing the original platform was reduced, the expression of RIPK1, p-RIPK1, NLRP3, cl-caspase-1 and ASC was up-regulated, and the contents of IL-1β and IL-18 were increased ( P<0.05), and pathological changes of hippocampal neurons were marked in PS group, DPS group and NPS group. Compared with PS group and DPS group, the escape latency was significantly shortened, the time of staying at the original platform quadrant was prolonged, the number of crossing the original platform was increased, the expression of RIPK1, p-RIPK1, NLRP3, cl-caspase-1 and ASC was down-regulated, the contents of IL-1β and IL-18 were decreased ( P<0.05), and pathological changes of hippocampal neurons were significantly attenuated in NPS group. Conclusions:Postoperative hippocampal RIPK1 function is enhanced in aged rats with chronic knee arthritis pain, which then activates NLRP3 inflammasomes, triggering neuroinflammation, and this process may be involved in the mechanism of postoperative neurocognitive dysfunction.

Objective To explore the effects of monocular deprivation(MD)on the expression of astrocytes in the superior colliculus,hippocampus,and visual cortex in mice during the critical period of visual development.Methods Eighteen C57BL/6J mice were randomly divided into the normal control group(CON group)and the MD group,with 9 mice in each group.Mice were bred under the 12 h/12 h dark/light conditions.Mice in the CON group received no treat-ment,while mice in the MD group underwent MD of the right eye on postnatal day 27,and the tissue was removed after 7 days.The mRNA and protein expression levels of glial fibrillary acidic protein(GFAP)in the superior colliculus,hippo-campus and visual cortex of mice in the two groups were detected using the real-time reverse transcription quantitative pol-ymerase chain reaction(RT-qPCR)and Western blot,respectively.The number of astrocytes labeled by GFAP and central nervous system specific protein β(S100β)in the superior colliculus,hippocampus and visual cortex of mice in the two groups was detected using the immunofluorescence staining.Results RT-qPCR and Western blot results showed that compared with the CON group,the mRNA and protein expression levels of GFAP in the superior colliculus,hippocampus(CA1,CA3 and dentate gyrus)and visual cortex of mice in the MD group decreased,and the differences were statistically significant(all P＜0.05).The immunofluorescence staining results showed that compared with the CON group,the number of GFAP and S1OOβ co-labeled astrocytes in the superior colliculus,hippocampus(CA1,CA3 and dentate gyrus)and visual cortex of mice in the MD group decreased,and the differences were statistically significant(all P＜0.05).Conclusion MD of mice during the critical period of visual development can result in a decrease in the number of astrocytes in the supe-rior colliculus,hippocampus and visual cortex.

Eclipta prostrata L.has been used in traditional medicine and known for its liver-protective properties for centuries.Wedelolactone(WEL)and demethylwedelolactone(DWEL)are the major coumarins found in E.prostrata L.However,the comprehensive characterization of these two compounds on non-alcoholic fatty liver disease(NAFLD)still remains to be explored.Utilizing a well-established zebrafish model of thioacetamide(TAA)-induced liver injury,the present study sought to investigate the impacts and mechanisms of WEL and DWEL on NAFLD through integrative spatial metabolomics with liver-specific transcriptomics analysis.Our results showed that WEL and DWEL significantly improved liver function and reduced the accumulation of fat in the liver.The biodistributions and metabolism of these two compounds in whole-body zebrafish were successfully mapped,and the discriminatory endogenous metabolites reversely regulated by WEL and DWEL treatments were also characterized.Based on spatial metabolomics and transcriptomics,we identified that steroid biosynthesis and fatty acid metabolism are mainly involved in the hepatoprotective effects of WEL instead of DWEL.Our study unveils the distinct mechanism of WEL and DWEL in ameliorating NAFLD,and presents a"multi-omics"platform of spatial metabolomics and liver-specific transcriptomics to develop highly effective compounds for further improved therapy.

Objective To investigate the effect of resveratrol(RES)on ovarian function in mice and elucidate the potential mechanism involving miR-23a.Methods Thirty mice were randomly divided into control,premature ovarian failure(POF)model,and treatment(RES)groups,with n=10 per group.The body weight and ovarian mass of the mice were measured,and the ovarian index was calculated.Hematoxylin and eosin staining was performed to observe pathological changes in mouse ovarian tissue.Real-time quantitative PCR and Western blotting were performed to measure the mRNA and protein levels of miR-23a,X-linked inhibitor of apoptosis protein(XIAP),and caspase-3 in the ovarian tissue.Results Compared with the control group,the POF group exhibited significant decreases in ovarian mass(P＜0.05),ovarian index(P＜0.05),number of primary follicles,and XIAP mRNA expression(P＜0.05),alongside significant increases in miR-23a and caspase-3 mRNA expression(P＜0.05).Compared with the POF group,the RES group exhibited significant increases in the ovarian mass(P＜0.05),ovarian index(P＜0.05),number of primary follicles,and XIAP mRNA expression(P＜0.05),as well as significant decreases in miR-23a and caspase-3 mRNA expression(P＜0.05).XIAP protein expression was significantly lower and caspase-3 protein expression was significantly higher in the POF group than in the control group(P＜0.05).Conversely,XIAP protein expression was significantly higher and caspase-3 protein expression was significantly lower in the RES group than in the POF group(P＜0.05).Conclusion RES exerts a protective effect on weakened ovarian function in mice,potentially mediated through its effect on miR-23a targeting the XIAP-caspase-3 signaling pathway.

Objective:To investigate the incidence rate and gene variation of methylmalonic academia (MMA) in Ji′nan city by analyzing biochemical and genetic screening results, and to explore the carrier frequency of MMA-related pathogenic genes in the population in Ji′nan.Methods:The children diagnosed with MMA by tandem mass spectrometry screening in Ji′nan Neonatal Disease Screening Centre from May 2011 to May 2022 were enrolled in this study.Their genetic test results were retrospectively analyzed and summarized.The dried heel blood tablets collected from 6 800 newborns were tested for neonatal gene screening. MMAA, MMAB, MMACHC and MMUT genes in 4 800 cases were detected by high-throughput sequencing+ target area capture technology.Ultra-multiplex polymerase chain reaction+ target gene locus capture technology was used to detect 174 target loci of 8 genes related to MMA in 2 000 cases.The hotspot mutation and related gene carrier rate of MMA were analyzed. Results:A total of 367 452 newborns were screened by tandem mass spectrometry, and 103 cases (56 males and 47 females) were diagnosed with MMA by screening.The estimated incidence of MMA was 1∶3 567.Among the 103 MMA cases, 76 were genetically diagnosed, and 4 gene variants of MMA ( MMAHC, MMUT, MMAA, MMADHC) were identified.A total of 6 800 neonates underwent neonatal genetic screening.Three of them were diagnosed with MMA.About 318 infants carried pathogenic variants of MMA, with a total carrier rate of 4.68%.Specifically, the carrier rates of MMACHC and MMUT gene variants were 3.09%(210/6 800) and 1.43% (97/6 800), respectively. Conclusions:MMA is the most common organic acid metabolism disorder in our country.The incidence and carrier rate of this disease are high in Jinan city.Neonatal genetic screening is an important supplement to neonatal biochemical screening.Carrier screening for MMA-related pathogenic genes is recommended for couples of childbearing age in Jinan.

Objective To investigate the expression of(glutathione S-Transferase Omega-1,GSTO1)in cervical cancer tissue and its correlation with patient survival time,and to explore the impact of GSTO1 N-glycosylation on proliferation,migration,invasion,and epithelial-mesenchymal transition of cervical cancer.Methods By using immunohistochemistry,the expression levels of GSTO1 in tumor cells of 82 cervical cancer patients were detected,and the correlation between GSTO1 expression and clinical pathological features was analyzed.Kaplan-Meier meth-od was used to plot survival curves and evaluate the impact of GSTO1 expression in cervical cancer tissues on pa-tient survival time.Univariate and multivariate Cox regression analyses were performed to assess the independent prognostic factors influencing cervical cancer prognosis.The NetNGlyc 1.0 Server database predicted potential N-glycosylation modification sites of GSTO1(Asn55,Asnl35,Asn190).The cervical cancer cells(HeLa)were transfected with GSTO1 N-glycosylation site mutation vectors at positions 55,135,and 190,as well as GSTO1 wild-type vector and empty vector.Stable transfected cells were selected using puromycin.Western blot experi-ments were performed to assess the effectiveness of lentiviral interference.The effects of GSTO1 N-glycosylation site mutations on proliferation,migration,and invasion of HeLa cells were evaluated using EdU proliferation assay,wound healing assay,and Transwell assay.The effect of GSTO1 N-glycosylation site mutations on the epithelial-mesenchymal transition of HeLa cells was detected using the Western blot experiment.Results Immunohistochem-istry results revealed high expression of GSTO1 in cervical cancer tissues.The expression rate of GSTO1 was signifi-cantly higher in cervical cancer tissues with deep stromal invision≥1/2,lymphovascular space invasion,and lymph node metastasis(P＜0.05).Moreover,high expression of GSTO1 was associated with poorer overall surviv-al.After N-glycosylation site-specific mutation of GSTO1,the cell count of proliferation,migration,and invasion in HeLa cells significantly decreased(P＜0.05).The Western blot results showed that N-glycosylation site mutation of GSTO1 significantly inhibited the epithelial-mesenchymal transition of HeLa cells.Conclusion The expression of GSTO1 in cervical cancer tissues is associated with stromal infiltration depth,lymphovascular space invasion and lymph node metastasis,and it is also correlated with shorter patient survival time.Site-specific mutations in GSTO1 N-glycosylation significantly inhibit the proliferation,migration and epitheli al-mesenchymal transition of HeLa cells.

OBJECTIVES: To retrospectively analyze the variation and characteristics of phenylalanine hydroxylase (PAH) gene, and to observe the long-term treatment effect and follow-up of newborns with PAH deficiency. METHODS: Clinical data, treatment and follow-up results of 198 patients with PAH deficiency diagnosed by newborn screening in Jinan from 1996 to 2021 were collected. The genetic analysis of 55 patients with PAH deficiency diagnosed by newborn screening in Jinan and 213 patients referred from the surrounding areas of Jinan were summarized. Gene variations were checked by a customized Panel gene detection method. Blood phenylalanine-concentration and physical development indicators including height and weight were regularly monitored. Intellectual development was assessed using a neuropsychological development scale for patients aged 0-6 years and academic performance, and brain injury in patients was assessed using brain magnetic resonance imaging. RESULTS: c.728G>A, c.158G>A, c.721C>T, c.1068C>A, c.611A>G variations were common in PAH gene. The genotype of c.158G>A variation is compound heterozygous variation, with mainly a mild hyperpheny-lalaninemia. 168 patients with PAH deficiency who were followed-up regularly had normal physical development without dwarfism or malnutrition. Among the 33 preschool patients who underwent mental development assessment, 2 were mentally retarded and the initial treatment age was older than 6 months. Nine patients with an average age of (17.13±2.42) years completed brain magnetic resonance imaging, one case was normal, and 8 cases were abnormal. There were patchy or patchy hyperintense foci near the bilateral lateral ventricles on T₂WI, and the intellectual development was normal. Compared with the other eight patients, the blood phenylalanine concentration of the normal child was better and stably controlled within the ideal range. CONCLUSIONS c.728G>A, c.158G>A, c.721C>T, c.1068C>A, c.611A>G variations were common in PAH gene. After standardized treatment, most patients with PAH deficiency diagnosed by screening can obtain normal growth and intellectual development in adolescence, but there are different degrees of organic lesions in the cerebral white matter.
Child ; Child, Preschool ; Adolescent ; Humans ; Infant, Newborn ; Young Adult ; Adult ; Neonatal Screening ; Follow-Up Studies ; Retrospective Studies ; Phenylketonurias/genetics* ; Phenylalanine Hydroxylase/genetics* ; Phenylalanine/therapeutic use* ; Mutation