Abstract： ObjectiveThe study aims to explore the effects and regulatory roles of glucose transporter 1 （GLUT1） on the proliferation， migration， adhesion， and angiogenesis of human umbilical vein endothelial cells （HUVECs） under ischemia-hypoxic conditions. MethodsIn vitro experiments were conducted to subject HUVECs to an ischemia-hypoxic-mimicking environment （1% O₂， 5% CO₂， 94% N₂）. The biological characteristics of HUVECs under normoxic and ischemia-hypoxic conditions were compared by assessing cell viability， proliferation capacity， and examining the expression changes of GLUT1， HIF-1α， and VEGFA proteins under ischemia-hypoxia using Western blot technology. Further， GLUT1 was overexpressed using plasmid transfection and the proliferation， migration， adhesion， and angiogenic capabilities of HUVECs were evaluated through scratch assays， cell adhesion assays， and tube formation assays. Mitochondrial morphological changes were observed by transmission electron microscopy，and oxygen consumption rate （OCR） was detected by Seahorse metabolic analyzer to evaluate mitochondrial function. ResultsCompared with normoxic conditions， the ischemia-hypoxic environment significantly inhibited the proliferation， cell viability， migration， and adhesion capabilities of HUVECs and impaired their angiogenic potential. The expression levels of GLUT1， HIF-1α and VEGFA proteins were also markedly reduced. However， when GLUT1 expression was upregulated， the migration， adhesion， and angiogenic capabilities of HUVECs were significantly improved， and the protein expression levels of HIF-1α， VEGFA and VEGFR were increased. Transmission electron microscopy revealed that ischemic-hypoxia leads to mitochondrial swelling and matrix damage， while GLUT1 overexpression significantly alleviates mitochondrial morphology abnormalities. OCR results suggest that GLUT1 overexpression may enhance oxidative phosphorylation of endothelial cells in ischemic-hypoxic environments to improve energy metabolism. These results suggest that GLUT1 may influence the function and angiogenic potential of HUVECs by regulating glucose metabolism and energy supply. ConclusionsThis study reveals the significant regulatory role of GLUT1 in the function of HUVECs under ischemia-hypoxic conditions， potentially through modulating cellular energy metabolism and signal transduction pathways， thereby affecting cell proliferation， migration， adhesion， and angiogenesis. These findings provide a new perspective on the role of GLUT1 in cardiovascular diseases and may offer potential targets for the development of new therapeutic strategies.

OBJECTIVE: To explore the improvement effect of electroacupuncture (EA) based on Xingnao Kaiqiao acupuncture (acupuncture for regaining consciousness and opening orifices) on cognitive impairment in mice with Parkinson's disease (PD), and to explore its regulatory mechanisms on the kinesin family member 5A (KIF5A)/mitochondrial Rho GTPase 1 (Miro1) pathway and mitophagy in prefrontal cortical neurons. METHODS: A total of 70 male C57BL/6J mice of clean grade were randomly divided into a normal group (12 mice), a sham operation group (12 mice), and a model pre-screening group (46 mice). Unilateral stereotaxic injection of 6-hydroxydopamine (6-OHDA) into the medial forebrain bundle was adopted to establish the PD model in the model pre-screening group. Twenty-four mice after successful modeling were randomly selected and divided into a model group and an EA group, 12 mice in each one. In the EA group, acupuncture was applied at "Shuigou" (GV26) and bilateral "Sanyinjiao" (SP6) and "Neiguan" (PC6), ipsilateral "Sanyinjiao" (SP6) and "Neiguan" (PC6) were connected to EA respectively, with disperse-dense wave, 5 Hz/20 Hz in frequency, 0.5 mA in current intensity, 20 min a time, 6 times a week for 30 days. Cognitive function was assessed by Y-maze and Morris water maze tests; morphology of prefrontal cortex was observed by H.E. staining; reactive oxygen species (ROS) level in prefrontal cortex was detected by fluorescence probe method; mitochondrial morphology and autophagosome ultrastructure were observed by transmission electron microscopy; the mRNA expression of tyrosine hydroxylase (TH) was detected by quantitative real-time PCR; the protein expression of TH, KIF5A, Miro1, p62, Parkin and PTEN induced kinase 1 (PINK1) was detected by Western blot. RESULTS: Compared with the sham operation group, both the model group and the EA group exhibited increased rotation number of per minute (P<0.001). Compared with the sham operation group, in the model group, the novel arm exploration time of Y-maze test was shortened (P<0.001), the escape latency of Morris water maze test was prolonged (P<0.05) and the platform crossing number of Morris water maze test was reduced (P<0.01); in the prefrontal cortex, the number of cellular vacuole and neurons with karyopyknosis was increased (P<0.001), and mitochondrial autophagosomes could be observed; in the prefrontal cortex, the relative expression of ROS was increased (P<0.001), the protein and mRNA expression of TH was decreased (P<0.001), the protein expression of Miro1, PINK1, Parkin was increased (P<0.001, P<0.01), the protein expression of KIF5A and p62 was decreased (P<0.001). Compared with the model group, in the EA group, the novel arm exploration time of Y-maze test was prolonged (P<0.01), the escape latency of Morris water maze test was shortened (P<0.05) and the platform crossing number of Morris water maze test was increased (P<0.05); in the prefrontal cortex, the number of cellular vacuole and neurons with karyopyknosis was decreased (P<0.001), and the number of mitochondrial autophagosomes reduced and the mitochondrial morphology was improved; in the prefrontal cortex, the relative expression of ROS was decreased (P<0.01), the protein and mRNA expression of TH was increased (P<0.001, P<0.01), the protein expression of Miro1, PINK1, Parkin was decreased (P<0.001, P<0.01, P<0.05), the protein expression of KIF5A and p62 was increased (P<0.01, P<0.05). CONCLUSION Xingnao Kaiqiao electroacupuncture effectively alleviates cognitive impairment and damage of neuronal function in PD mice, its mechanism may be related to the regulation of KIF5A/Miro1 pathway, hence reducing the mitophagy in prefrontal cortical neurons.
Animals ; Electroacupuncture ; Male ; Mice ; Parkinson Disease/physiopathology* ; Cognitive Dysfunction/psychology* ; Kinesins/genetics* ; Humans ; Mitophagy ; Mice, Inbred C57BL ; rho GTP-Binding Proteins/genetics* ; Mitochondria/genetics* ; Prefrontal Cortex/metabolism*

Objective To investigate prokaryotic expression of the antigen sequence (amino acids 59-145) of mouse leukemia-related protein 16 (LRP16) protein and preparation of rabbit anti-mouse LRP16 polyclonal antibody. Methods The prokaryotic expression plasmid pLS962-LRP16 was constructed by the molecular cloning method and transferred into E.coli Rosetta to express LRP16 protein induced by IPTG. The recombinant protein was purified using Ni-NTA affinity columns followed by gel filtration chromatography. New Zealand white rabbits were immunized with the purified antigen to generate polyclonal antiserum, with antibody titer quantified by ELISA. Antigen-specific IgG was affinity-purified using Sepharose-coupled LRP16 and validated through Western blot and immunofluorescence assays. Results SDS-PAGE analysis confirmed insoluble expression of the LRP16 fusion protein as inclusion bodies. ELISA demonstrated exceptional antiserum titer (1:256 000). Western blot and immunofluorescence verified that the polyclonal antibody could specifically recognize endogenous LRP16 in murine tissues. Conclusion The prokaryotic expression of the LRP16 gene is successfully achieved, and the rabbit anti-mouse LRP16 polyclonal antibody exhibiting high specificity is prepared. This lays the foundation for further studies on the function of the LRP16 gene.
Animals ; Rabbits ; Mice ; Antibodies/immunology* ; Escherichia coli/metabolism* ; Enzyme-Linked Immunosorbent Assay ; Blotting, Western ; Antibody Specificity

Skeletal muscle dysfunction is a common extrapulmonary comorbidity of chronic obstructive pulmonary disease (COPD) and is associated with decreased quality-of-life and survival in patients. The autophagy lysosome pathway is one of the proteolytic systems that significantly affect skeletal muscle structure and function. Intriguingly, both promoting and inhibiting autophagy have been observed to improve COPD skeletal muscle dysfunction, yet the mechanism is unclear. This paper first reviewed the effects of macroautophagy and mitophagy on the structure and function of skeletal muscle in COPD, and then explored the mechanism of autophagy mediating the dysfunction of skeletal muscle in COPD. The results showed that macroautophagy- and mitophagy-related proteins were significantly increased in COPD skeletal muscle. Promoting macroautophagy in COPD improves myogenesis and replication capacity of muscle satellite cells, while inhibiting macroautophagy in COPD myotubes increases their diameters. Mitophagy helps to maintain mitochondrial homeostasis by removing impaired mitochondria in COPD. Autophagy is a promising target for improving COPD skeletal muscle dysfunction, and further research should be conducted to elucidate the specific mechanisms by which autophagy mediates COPD skeletal muscle dysfunction, with the aim of enhancing our understanding in this field.
Pulmonary Disease, Chronic Obstructive/physiopathology* ; Autophagy/physiology* ; Humans ; Muscle, Skeletal/pathology* ; Mitophagy ; Animals ; Mitochondria/metabolism* ; Lysosomes

A 52-year-old female patient was diagnosed with eosinophilic granulomatosis with polyangiitis (EGPA), asthma and allergic rhinitis and was treated with mepolizumab (100 mg, once every 4 weeks). Three days after the second administration, the patient developed dark yellow urine, and abnormal liver function indicators were found on day 6 after administration. Laboratory tests showed total bilirubin (TBIL) of 22 μmol/L, direct bilirubin (DBIL) of 14.9 μmol/L, alanine aminotransferase (ALT) of 461 U/L, aspartate aminotransferase (AST) of 129 U/L, and alkaline phosphatase (ALP) of 296 U/L. After a comprehensive assessment, it was considered that mepolizumab may caused liver injury in the patient. Magnesium isoglycyrrhizinate, glutathione, and ursodeoxycholic acid were used for liver protection. Five days later, the urine color became lighter and liver function indicators improved (TBIL 17.4 μmol/L, DBIL 9.8 μmol/L, ALT 214 U/L, AST 85 U/L, ALP 226 U/L). After about 2 weeks of continued liver protection treatment, liver function returned to normal. Mepolizumab treatment was suspended and during the follow-up for about half a year, the patient did not experience liver function abnormalities or other discomforts again.

Objective:To evaluate the effect of sodium sivelestat on the expression of specialized pro-resolving mediators (SPMs) synthesis enzymes in mice with lipopolysaccharide (LPS)-induced acute lung injury (ALI).Methods:Eighteen SPF healthy male C57BL/6 mice, aged 6-8 weeks, weighing 20-25 g, were divided into 3 groups ( n=6 each) using a random number table method: control group (C group), LPS-induced ALI group (ALI group), and LPS-induced ALI + sodium sivelestat group (ALI+ SV group). ALI was induced by intravenous injection of LPS 15 mg/kg through the tail vein. Sodium sivelestat 50 mg/kg was intraperitoneally injected at 1 h after LPS administration. At 12 h after LPS administration, blood samples were collected from the eyeballs for routine blood tests, and the remaining blood was processed for serum extraction. The mice were sacrificed after anesthesia, and lung tissues were collected to determine the wet/dry weight (W/D) ratio, serum concentrations of interleukin-1beta (IL-1β) and IL-10 (by enzyme-linked immunosorbent assay), expression of neutrophil elastase (NE) and SPMs synthesis enzymes 5-lipoxygenase (5-LOX), 12-lipoxygenase (12-LOX), and 15-lipoxygenase (15-LOX) in lung tissues (by Western blot) and to examine the pathological changes of lung tissues which were scored. Results:Compared with C group, the lung injury scores, W/D ratio, white blood cell counts, percentage of neutrophil, and serum IL-1β and IL-10 concentrations were significantly increased, the expression of NE was up-regulated, and the expression of 5-LOX, 12-LOX and 15-LOX was down-regulated in ALI group ( P<0.05). Compared with ALI group, the lung injury scores, W/D ratio, white blood cell counts, percentage of neutrophil, and serum IL-1β concentration were significantly decreased, the serum IL-10 concentration was increased, the expression of NE was down-regulated, and the expression of 5-LOX, 12-LOX and 15-LOX was up-regulated in ALI+ SV group ( P<0.05). Conclusions:The mechanism by which sodium sivelestat alleviates LPS-induced ALI may be related to up-regulating the expression of SPMs synthesis enzyme and promoting the resolution of pulmonary inflammation in mice.

Objective To explore the correlation between serum residual lipoprotein cholesterol(RLP-C)and triglyceride glucose(TyG)index in postmenopausal type 2 diabetes(T2DM)patients.Methods A total of 389 postmenopausal T2DM patients who attended Ruili People's Hospital from May 2019 to May 2023 were included.RLP-C and TyG index were calculated,and the study subjects were divided into the low TyG index group(n=194)and the high TyG index group(n=195)based on the median of TyG index(9.23).Differences in general clinical data and metabolism-related indexes between the two groups of patients were compared,and Spearman rank correlation was performed to explore their correlation.Single-factor and multivariate Logistic regressions were used to explore their influencing factors,and ROC curve was used to analyze their diagnostic value.Results Compared with the low TyG index group,the history of hypertension(79.49%vs 70.62%),hyperlipidemia(22.05%vs 13.40%),hemoglobin(Hb)(120.34±19.96g/L vs 114.97±21.32g/L),fasting blood glucose(FBG)[3.97(3.03,5.10)mmol/L vs 3.64(2.99,4.74)mmol/L],total cholesterol(TC)[5.00(4.40,5.95)mmol/L vs 4.36(3.78,5.30)mmol/L],triacylglycerol(TG)[2.11(1.60,3.00)mmol/L vs 1.20(0.91,1.54)mmol/L],low-density lipoprotein(LDL-C)[[2.99(2.43,3.93)mmol/L vs 2.71(2.13,3.38)mmol/L],non-high-density lipoprotein(nonHDL-C)[3.94(3.22,4.82)mmol/L vs 3.15(2.53,3.94)mmol/L]and RLP-C concentration[0.76(0.52,1.08)mmol/L vs 0.44(0.29,0.59)mmol/L]of the high TyG index group were higher,while high-density lipoprotein(HDL-C)concentration was lower,and the differences were statistically significant(x2=4.09,4.99;t=-2.56;Z=-2.34,-5.15,-12.08,-3.04,-6.23,-9.15,-3.99,all P<0.05).Spearman correlation analysis showed that TyG in all samples was positively correlated with TC,TG,LDL-C,RLP-C and nonHDL-C(r=0.304,0.769,0.179,0.386,0.571,all P<0.001),but was negatively correlated with HDL-C(r=-0.306,P<0.001).The correlations between TyG and TC,LDL-C in the high TyG index group were not significant(all P>0.05).Single-actor logistic regression analysis showed that history of hypertension,hyperlipidemia,HDL-C,LDL-C,and RLP-C were the factors influencing TyG levels[OR(95%CI)=1.61(1.01～2.57),1.83(1.07～3.12),0.28(0.14～0.54),1.21(1.02～1.43),17.58(8.11～38.11),all P<0.05],while the multivariate regression analysis showed that only RLP-C was an independent risk factor for elevated TyG[OR(95%CI)=13.17(5.71～30.37),P<0.001).ROC curves showed that the AUC(95%CI)for the diagnosis of TyG index elevation by RLP-C was 0.768(95%CI:0.721～0.816),with the cutoffvalue of 0.59 mmol/L.The sensitivity and specificity were 69.71%and 75.77%,respectively.Conclusion RLP-C is an independent risk factor for elevated TyG in postmenopausal T2DM patients,which has some clinical value in predicting insulin resistance.

【Objective】 To develop the Visual Cognitive Ability Assessment Scale for Preschool Children and to evaluate its reliability and validity, in order to provide reference for clinical evaluation of visual cognitive ability of preschool children. 【Methods】 1) From November 2021 to February 2022, construct the dimension framework was constructed and the pool of scale items was compiled according to the theory. 2) In March to June 2022, items were screened preliminarily through group discussion and Delphi method. In August 2022, the entries was revised by a pre-survey in a small sample (n=50). 3) Parents of children aged 4 - 7 from 8 kindergartens in 4 main urban areas of Nanjing were investigated by stratified cluster random sampling method in September to December 2022. The valid sample of the first survey (n=344) was analyzed to conduct item analysis and re-test reliability analysis, the valid sample of the second survey (n=695) was tested for reliability and confirmatory factor analysis, then the scale was finally compiled and evaluated. 【Results】 1) The scale contained 19 items in 4 dimensions:visual memory, discerning vision, spatial vision and visual integration. All items passed the project analysis test. 2) The Cronbach′s α coefficient of each dimension ranged from 0.604 to 0.886, and the Cronbach′s α coefficient of the whole scale was 0.917. During the two surveys, the scores of each item were correlated, and the average retest reliability coefficient was 0.601 (P< 0.05). 3) Content validity index (S-CVI) at scale level was 0.91, and item level content validity index (I-CVI) ranged from 0.8 to 1.0. After several rounds of model modification, the confirmatory factor model fit well. 【Conclusion】 The reliability and validity of the Visual Cognitive Ability Assessment Scale for Preschool Children are acceptable and meet the requirements of the scale, which provides a practical tool for clinical screening of visual and cognitive disorders.

The methodological framework for interventional clinical research of Chinese medicine （CM） under the research mode of syndrome dominating disease provides a set of technical principles and methods to design， evaluate， and implement of this kind. It consists of three main parts including general principles， research points and key design elements， with a total of 25 items. This methodological framework proposes implementing requirements and recommendations in a variety of aspects， including basic norms to be followed in relevant studies， perspectives for selecting research topics， as well as the technological details on study population （P）， intervention （I） and comparison（C）， outcome measurement （O）， time frame （T） of treatment and follow-up， sample orientation （prospective versus retrospective）， study design （S） format and type. To provide practical guidance for future studies， this article clearly explains each items of the methodological framework through some supportive cases.

Objective:To observe the therapeutic effect of intraocular lens (IOL) protected phacoemulsification (PHACO) in patients with hard nucleus cataract.Methods:A randomized controlled clinical study was conducted.A total of consecutive 120 patients (120 eyes) with hard nucleus cataract of Emery grade Ⅳ or Ⅴ were enrolled from January 2019 to May 2022.The patients were randomly divided into PHACO group receiving routine PHACO, IOL protected PHACO group receiving PHACO under IOL protection, and extracapsular cataract extraction (ECCE) group receiving ECCE, with 40 cases (40 eyes) in each group.Finally, 99 patients completed the follow-up, including 30 cases (30 eyes) in PHACO group, 35 cases (35 eyes) in IOL protected PHACO group, and 34 cases (34 eyes) in ECCE group.The total operation time, intraoperative PHACO time and cumulative energy release of each patient were recorded.The corneal endothelial cell density (ECD), coefficient of variation in endothelial cell area (CV), hexagonal endothelial cell ratio (6A), corneal astigmatism and the number of eyes with different grades of uncorrected visual acuity were measured and compared after 3-month follow-up.The intraoperative and postoperative complications were recorded.This study adhered to the Declaration of Helsinki and was approved by the Ethics Committee of Yanbian University Hospital (NO.2023002).Patients were informed of study content and purpose and signed a consent form before treatment.Results:There was no significant difference in ultrasonic energy and time between PHACO group and IOL protected PHACO group ( P=0.691, 0.982).The total operation time was (38.81±2.73) and (36.45±3.45) minutes in PHACO group and IOL protected PHACO group, significantly shorter than (69.60±4.35) minutes in ECCE group (both at P<0.001).There was no significant difference in age, sex, lens nucleus hardness and other baseline data among the three groups before operation (all at P>0.05).Three months after operation, the number of patients with higher uncorrected visual acuity in PHACO group and IOL protected PHACO group was larger than that in ECCE group ( P=0.006, 0.007).The ECD and 6A in IOL protected PHACO group were (2 155.57±177.88)/mm 2 and (41.31±5.18)%, respectively, which were significantly higher than (1 912.64±224.11)/mm 2 and (36.18±3.27)% in PHACO group, and the CV in IOL protected PHACO group was (50.34±5.90)%, which was lower than (55.67±3.30)% in PHACO group, showing statistically significant differences ( P=0.007, 0.003, 0.005).At 1 week and 3 months after the operation, the corneal astigmatism was significantly lower in IOL-protected PHACO group than in ECCE group, but higher than in PHACO group, and the difference were statistically significant (all at P<0.05). Conclusions:Compared with conventional PHACO, IOL-protected PHACO can effectively reduce the damage of corneal endothelium caused by ultrasonic energy, shorten the operation time and reduce postoperative inflammatory reaction compared with ECCE, and does not significantly increase postoperative corneal astigmatism.IOL-protected PHACO is an effective improved surgical method for patients with hard nucleus cataract.