OBJECTIVE: To investigate the clinical characteristics, steroid hormone profiles, and genetic variants in two female patients with Non-classic adrenal hyperplasia (NCAH). METHODS: Clinical data and samples were collected from two patients who had visited Huaian Maternal and Child Health Care Hospital Affiliated to Medical College of Yangzhou University on September 27, 2022 and June 25, 2023, respectively, with an initial diagnosis of Polycystic ovary syndrome (PCOS) and suspected NCAH. Seven steroid hormones in dried blood spots were analyzed using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Single base variants and repeat/deletions in the CYP21A2 gene were analyzed by using a classic congenital adrenal hyperplasia (CAH) gene assay, and 10 related genes were analyzed by third-generation sequencing (TGS) should the variants be unclear. This study has been approved by the Medical Ethics Committee of the hospital (Ethics No.: 2025003). RESULTS: Patient 1 was a 14-year-old girl, and patient 2 was a 23-year-old woman with insulin resistance. Both patients had hirsutism, acne, bilateral polycystic ovarian morphology, in addition with significantly elevated serum testosterone by chemiluminescence. The steroid hormone profiles of both patients suggested a significant increase in 17-hydroxyproesterone, normal cortisol and 11-deoxycortisol. Patient 2 additionally showed a significant rise in 21-deoxycortisol. The presentation of both patients was indicative of NCAH, which was also evidenced by their respective medical histories. Sanger sequencing of long fragment PCR amplification combined with multiplex ligation-dependent probe amplification (MLPA) revealed that patient 1 harbored a mild c.92C>T (p.P31L) variant and a severe variant with a large segmental deletion in CYP21A2. Patient 2 was finally confirmed by TGS to carry mild CYP21A2 variants in the 5' untranslated region (5' UTR) promotor region (c.-126C>T, c.-113G>A, c.-110T>C) and a severe c.293-13C/A>G variant. The promotor region variants had resulted in decompression of the long fragment P1X/P2 amplification, leading to homozygous result of Sanger sequencing for c.293-13C/A>G, which in turn halved the amplification signal for the wt-113 SNP probe. In addition, the wtI2G-A probe was enhanced by interference in the MLPA assay. CONCLUSION This study demonstrated that NCAH should be excluded when PCOS is accompanied by a significant increase in serum testosterone, that mass spectrometry of steroid hormone profiles containing 17-hydroxyprogesterone is useful for the detection of NCAH, and that TGS is advantageous in confirming the diagnosis of NCAH when compared with conventional genetic testing methods.
Humans ; Female ; Adrenal Hyperplasia, Congenital/blood* ; Adolescent ; Steroid 21-Hydroxylase/genetics* ; Young Adult ; Genetic Variation ; Adult

Subarachnoid hemorrhage (SAH) is a serious intracranial hemorrhage characterized by acute bleeding into the subarachnoid space. The effects of shikonin, a natural compound from the roots of Lithospermum erythrorhizon, on oxidative stress and blood–brain barrier (BBB) injury in SAH was evaluated in this study. A rat model of SAH was established by endovascular perforation to mimic the rupture of intracranial aneurysms. Rats were then administered 25 mg/kg of shikonin or dimethylsulfoxide after surgery. Brain edema, SAH grade, and neurobehavioral scores were measured after 24 h of SAH to evaluate neurological impairment. Concentrations of the oxidative stress markers superoxide dismutase (SOD), glutathione (GSH), and malondialdehyde (MDA) in the brain cortex were determined using the corresponding commercially available assay kits. Evans blue staining was used to determine BBB permeability. Western blotting was used to quantify protein levels of tight junction proteins zonula occludens-1, Occludin, and Claudin-5. After modeling, the brain water content increased significantly whereas the neurobehavioral scores of rats with SAH decreased prominently. MDA levels increased and the levels of the antioxidant enzymes GSH and SOD decreased after SAH. These changes were reversed after shikonin administration. Shikonin treatment also inhibited Evans blue extravasation after SAH. Furthermore, reduction in the levels of tight junction proteins after SAH modeling was rescued after shikonin treatment. In conclusion, shikonin exerts a neuroprotective effect after SAH by mitigating BBB injury and inhibiting oxidative stress in the cerebral cortex.

Purpose To quantitatively compare the diffusion parameters of mono-and biexponential diffusion-weighted imaging models,and to obtain optimal sets of b-values in diffusion-weighted MRI for obtaining monoexponential apparent diffusion coefficient(ADC)close to perfusion-insensitive intravoxel incoherent motion(IVIM)model ADC(ADCIVIM)in identifying of pulmonary solid benign and malignant lesions.Materials and Methods IVIM was performed in 40 patients with solid nodule and masse in Xi'an Gaoxin Hospital from July 2021 to August 2022 using a 3.0T MR imager.Two experienced diagnostic radiologists subjectively evaluated the IVIM images.A single index model was used to calculate ADC values(ADC0-1 000,ADC20-1 000,ADC50-1 000,ADC80-1 000,ADC150-1 000,ADC300-1 000,ADC500-1 000,ADC300,500,1 000,ADC300,800,1 000,ADC300,500,ADC300,800 and ADC300,1 000).The reference standard ADCIVIM value were calculated using a double-exponential model.The physician's measurements between two physicians were measured.The malignant and benign groups were compared and receiver operator characteristic curve for all parameters were analyzed.Results The measurement consistency of ADC values under b value sets and ADCIVIM was very good,and the intraclass correlation coefficient was more significant than 0.75.The differences between ADCIVIM and ADC values in each b group were statistically significant(t=-6.016--2.500,all P<0.05).The area under the curve(AUC)of ADCIVIM was the largest(0.906),with an optimal threshold of 1.271×10-3 mm2/s,a sensitivity of 80.0%and a specificity of 93.0%.The diagnostic efficacy close to ADCIVIM were ADC300,800(AUC=0.891),ADC50-1 000(AUC=0.827)and ADC300,800,1 000(AUC=0.795),respectively.The optimal threshold of ADC300,800 was 1.140×10-3 mm2/s,the sensitivity and specificity were 80.0%and 93.7%,respectively.Conclusion Combining b-values 300 s/mm2 and 800 s/mm2 is recommended as routine scanning parameters for identifying the insensitive monoexponential ADC between benign and malignant solid pulmonary lesions.

Objective To explore the possible mechanism of T cell immunoglobulin and mucin domain-containing protein-3(TIM-3)regulating M2 macrophage polarization through Galectin-3 in lung injury induced by Streptococcals pneumoniae pneumonia(SPP)in mice.Methods Thirty mice were divided into control group,SPP group and SPP+TIM-3 inhibition group,with 10 mice in each group.The macrophage cell line RAW 264.7 was cultured and divided into a control group,with the SPP group versus the SPP+TIM-3 inhibition group.Lung histopathology was detected by hematoxylin-eosin(H&E)staining.Serum and cell supernatant tumor necrosis factor-α(TNF-α),interleukin-1β(IL-1β)and IL-6 content were measured by enzyme-linked immunosorbent assay(ELISA).Western blotting the protein levels of TIM-3 and Galectin-3 were determined in lung tissues and macrophages.Cluster of differentiation(CD)206,CD86 mRNA levels in lung tissue and macrophages were determined by real time quantitative PCR(RT-qPCR).Results Compared with the control group,the SPP group had significantly abnormal lung tissue pathological structure,thickened alveolar septum,a large number of inflammatory cells infiltration,a large amount of exudate in alveolar space,and increased serum inflammatory factors TNF-α,IL-6 and IL-1β,the protein and mRNA expressions of CD86,the marker protein of M1 macrophages,and the protein and mRNA expressions of CD206,the marker protein of M2 macrophages,and the protein expressions of TIM-3 and Galectin-3 in lung tissues were significantly increased in the model group,and the differences were statistically significally(t=8.36～76.30,all P<0.05).Compared with the SPP group,the SPP+TIM-3 inhibition group had improved lung tissue pathological damage,reduced alveolar septum thickness,inflammatory cell infiltration,reduced exudate in alveolar space,decreased serum inflammatory factors TNF-α,IL-6 and IL-1β content,and decreased expression of CD86 protein and mRNA in lung tissue.The expression of CD206 protein and mRNA was increased,the expression of TIM-3 protein was decreased,and the expression of Galectin-3 protein was increased,and the differences were statistically significant(t=10.67～63.32,all P<0.05).Cell experiments were consistent with this result.Conclusion Inhibition of TIM-3 can improve lung injury in SPP mice by down-regulating the expression of Galectin-3 and promoting M2 macrophage polarization.

Objective:To investigate the protective effect of achyranthes bidentata(AB)on sperm quality in mice with sper-matogenic disorder through the glycolytic metabolic pathway and its action mechanism.Methods:We equally randomized 40 Kun-ming mice into a normal control,a model control,a low-dose AB(3.5 g/kg)and a high-dose AB group(7.0 g/kg),and established the model of spermatogenic disorder in the latter three groups of mice by intraperitoneal injection of doxorubicin(30 mg/kg).Two days after modeling,we collected the testis and kidney tissues and blood samples from the mice for observation of the pathological changes in the testis tissue by HE staining,detection of perm motility with the sperm quality analyzer,examination of the apoptosis of testis cells by flow cytometry,measurement of the levels of testosterone(T),malondialdehyde(MDA),superoxide dismutase(SOD)and cata-lase(CAT)in the serum and testis tissue by ELISA,and determination of expressions of the key enzymes of glycolysis hexokinase Ⅱ(HK2),pyruvate kinase M2(PKM2),platelet phosphofructokinase(PFKP),lactate dehydrogenase A(LDHA)and the meiosis pro-teins REC8 and SCP3 by Western blot,and the mRNA expressions of glycolytic phosphofructokinase 1(PFK1),phosphoglycerate ki-nase 1(PGK1),tumor necrosis factor-α(TNF-α)and interleukin-1 β(IL-1β)by fluorescence quantitative PCR(FQ-PCR).Results:Compared with the model controls,the mice in the AB groups showed significant increases in the testis coefficient,kidney in-dex,sperm concentration,sperm motility,spermatogonia,primary spermatocytes,spermatids,sperm count and the serum T level(P＜0.05 orP＜0.01),but dramatic decreases in the apoptosis of testis cells and percentage of morphologically abnormal sperm(P＜0.01).Achyranthes bidentata also significantly elevated the levels of SOD and CAT,and down-regulated the mRNA expressions of MDA,TNF-α and IL-1β(P＜0.05 or P＜0.01),and up-regulated the protein expressions of HK2,PKM2,PFKP,LDHA,REC8 and SCP3,and expressions of the glycolysis key genes Pfk1 and Pgk1(P＜0.05 orP＜0.01).Conclusion:Achyranthes bidentata ameliorates doxorubicin-induced spermatogenic disorder in mice by regulating the glycolytic pathway and reducing oxidative stress and the expressions of inflammatory factors.

Aim To investigate the effects of Agrimonia pilosa(AP)on the proliferation and apoptosis of non-small cell lung cancer(NSCLC)H1299 cells using non-targeted metabolomics and other methods,and to explore the underlying molecular mechanisms.Meth-ods Taking H1299 cells as the research object,the effect of AP on cell proliferation and apoptosis was de-tected through CCK-8 method,colony formation,LDH,Hoechst 33258 staining,AO/EB staining,flow cytometry detection,RT qPCR and other experiments.The main differential metabolites were detected by the metabolomics method of ultra-high phase liquid chro-matography and mass spectrometry(UHPLC-Q-Orbi-trap MS),and related metabolic pathways were ana-lyzed.Results Compared with the control group,AP treatment was able to significantly inhibit the prolifera-tion and colony formation of H1299 cells,while the re-lease of LDH increased in a dose-dependent manner.Fluorescence microscopy and flow cytometry and RT-qPCR analysis revealed that H1299 cells underwent crumpling and increased nuclear fragmentation after AP administration,blocked in G0/G1 phase,up-regulated apoptotic genes caspase-3 and Bax,and down-regulated apoptosis-inducing effects of Bcl-2.Metabolomics anal-ysis screened 35 differential metabolites,which were PC(O-30∶1),D-Glutamic acid,PE(18∶0/15∶0),etc.The main metabolic pathways involved includ-ed amino acid metabolism,glycerophospholipid metabo-lism and purine metabolism so on.Conclusions AP may exert its pharmacological effects by interfering with multiple metabolic pathways in H1299 cells,inhibiting cell proliferation and promoting apoptosis.

Objective To investigate the clinical effect of shoulder arthroscopic rotator cuff repair combined with shoulder three needles in treatment of rotator cuff injury.Methods 86 patients with rotator cuff injury from June 2021 to May 2023 were divided into two groups using a random number table method.The research group received 43 cases of rotator cuff repair under arthroscopy combined with shoulder three needles rotator cuff repair,while the control group received 43 cases of rotator cuff repair under arthroscopy combined with placebo needling.Then compare the pain level,shoulder joint function,and shoulder joint range of motion between the two groups before and 6 months after surgery,and calculate the excellent and good shoulder joint function rates between the two groups at 6 months after surgery,and compare the changes in inflammatory factors between the two groups at before and 4 weeks after surgery.Results 6 months after surgery,the visual analogue scale(VAS)score of both groups was lower than that before surgery,and the VAS score in research group(including the moderate tearing subgroup and the major tearing subgroup)was lower than that of the control group(including the moderate tearing subgroup and the major tearing subgroup),while the American Shoulder and Elbow Surgeons(ASES)score and shoulder joint range of motion were higher than those before surgery,and the ASES score and shoulder joint range of motion in research group(including the moderate tearing subgroup and the major tearing subgroup)were higher than those of the control group(including the moderate tearing subgroup and the major tearing subgroup),the differences were statistically significant(P＜0.05).6 months after surgery,the excellent and good rate of shoulder joint function in the research group was higher than that in the control group,the difference was statistically significant(P＜0.05).At 4 weeks after surgery,the levels of tumor necrosis factor-α(TNF-α),C-reactive protein(CRP),interleukin-6(IL-6),and interleukin-8(IL-8)in both groups were lower than those before surgery,and the research group(including the moderate tearing subgroup and the major tearing subgroup)were lower than those of control group(including the moderate tearing subgroup and the major tearing subgroup),the differences were statistically significant(P＜0.05).Conclusion Arthroscopic rotator cuff repair combined with shoulder three needles can alleviate the pain level of patients with rotator cuff injury,improve shoulder joint function and range of motion,increase the excellent and good rate of shoulder joint function,and reduce inflammatory reaction.It is worthy of clinical application.

Objective To investigate the clinical effect of shoulder arthroscopic rotator cuff repair combined with shoulder three needles in treatment of rotator cuff injury.Methods 86 patients with rotator cuff injury from June 2021 to May 2023 were divided into two groups using a random number table method.The research group received 43 cases of rotator cuff repair under arthroscopy combined with shoulder three needles rotator cuff repair,while the control group received 43 cases of rotator cuff repair under arthroscopy combined with placebo needling.Then compare the pain level,shoulder joint function,and shoulder joint range of motion between the two groups before and 6 months after surgery,and calculate the excellent and good shoulder joint function rates between the two groups at 6 months after surgery,and compare the changes in inflammatory factors between the two groups at before and 4 weeks after surgery.Results 6 months after surgery,the visual analogue scale(VAS)score of both groups was lower than that before surgery,and the VAS score in research group(including the moderate tearing subgroup and the major tearing subgroup)was lower than that of the control group(including the moderate tearing subgroup and the major tearing subgroup),while the American Shoulder and Elbow Surgeons(ASES)score and shoulder joint range of motion were higher than those before surgery,and the ASES score and shoulder joint range of motion in research group(including the moderate tearing subgroup and the major tearing subgroup)were higher than those of the control group(including the moderate tearing subgroup and the major tearing subgroup),the differences were statistically significant(P＜0.05).6 months after surgery,the excellent and good rate of shoulder joint function in the research group was higher than that in the control group,the difference was statistically significant(P＜0.05).At 4 weeks after surgery,the levels of tumor necrosis factor-α(TNF-α),C-reactive protein(CRP),interleukin-6(IL-6),and interleukin-8(IL-8)in both groups were lower than those before surgery,and the research group(including the moderate tearing subgroup and the major tearing subgroup)were lower than those of control group(including the moderate tearing subgroup and the major tearing subgroup),the differences were statistically significant(P＜0.05).Conclusion Arthroscopic rotator cuff repair combined with shoulder three needles can alleviate the pain level of patients with rotator cuff injury,improve shoulder joint function and range of motion,increase the excellent and good rate of shoulder joint function,and reduce inflammatory reaction.It is worthy of clinical application.

Fetal malformation of cortical development(MCD)is a group of structural neurological disorders caused by abnormalities in development of cortical layer during embryogenesis,characterized by significant heterogeneity and diversity,which may lead to adverse clinical outcomes such as epilepsy and intellectual disabilities.The progresses in prenatal evaluation on fetal MCD were reviewed in this article.

Objective To evaluate the efficacy and safety of a novel intravascular lithotripsy(IVL)balloon—Vesscrack shockwave balloon—for vascular preparation before stent implantation in patients with severe coronary artery calcification(CAC).Methods This was a prospective,single-arm,multicenter study conducted in China from June 2022 to October 2022.Patients with severe CAC were treated with the Vesscrack shockwave balloon for lesion preparation,followed by drug-eluting stent(DES)implantation.Of these,33 patients underwent optical coherence tomography(OCT).The primary endpoint was procedural success,defined as successful stent implantation with residual stenosis≤30％and the absence of in-hospital major adverse events,including cardiac death,target vessel-related myocardial infarction,or target lesion revascularization.Results A total of 170 patients[mean age:(65.9±7.9)years,116 males]were enrolled.After treatment with IVL and DES,the minimum lumen diameter increased significantly compared to baseline[(2.34±0.40)mm vs.(0.95±0.33)mm,P＜0.001],the degree of stenosis was significantly reduced[(13.24±6.60)％vs.(65.18±10.59)％,P＜0.001].Procedural success was achieved in 100％of cases,and device success was 98.8％.The 30-day patient-related cardiovascular clinical composite endpoint(POCE)rate was 0.0,with no target lesion failure,no confirmed or potential thrombotic events were observed.The shockwave energy generator demonstrated excellent stability and ease of use.Among the 33 patients assessed with OCT,after IVL intervention,the maximum calcified area of the lumen[(3.51±1.51)mm2 vs.(2.85±1.80)mm2,P＜0.001],and the minimum lumen area within the target lesion[(3.08±1.04)mm2 vs.(2.02±0.75)mm2,P＜0.001],and after DES intervention,the luminal area of the largest calcified site[(6.59±1.64)mm2 vs.(2.85±1.80)mm2,P＜0.001]and the minimum luminal area within the target lesion[(6.19±1.45)mm2 vs.(2.02±0.75)mm2,P＜0.001]were significantly increased,and the differences were statistically significant.Conclusions The Vesscrack shockwave balloon is effective and safe for vascular preparation in patients with severe CAC prior to stent implantation.It achieves significant calcified plaque modification,high procedural success rates,and minimal complications.