miR-33-5p knockdown attenuates abdominal aortic aneurysm progression via promoting target adenosine triphosphate-binding cassette transporter A1 expression and activating the PI3K/Akt signaling pathway.

PURPOSE: The aim of this study was to investigate the role of miR-33-5p in abdominal aortic aneurysm progression, which regulated adenosine triphosphate-binding cassette transporter A1 (ABCA1)-mediated cholesterol efflux and lipid accumulation in THP-1 macrophage-derived foam cells through the PI3K/Akt pathway. METHODS: Quantitative reverse transcription polymerase chain reaction was used to evaluate the expression level of miR-33-5p and ABCA1 mRNA in abdominal aortic aneurysm patient and normal person tissues. The relationship between miR-33-5p and ABCA1 was examined by dual luciferase report assay. High-performance liquid chromatography was used to evaluate the levels of cholesterol contents. Cholesterol efflux detection was performed by liquid scintillator. The expression of inflammatory cytokines was detected by quantitative reverse transcription polymerase chain reaction. Western blot was applied to determine the expression levels of ABCA1, PI3K (p-PI3K), and Akt (p-Akt). RESULTS: The quantitative reverse transcription polymerase chain reaction analysis results revealed miR-33-5p overexpression in abdominal aortic aneurysm tissues, but the expression level of ABCA1 was lower in abdominal aortic aneurysm tissues than non-abdominal aortic aneurysm tissues. Subsequently, the dual luciferase report gene assay confirmed that ABCA1 was a target of miR-33-5p, and miR-33-5p-negative regulated ABCA1 expression. Moreover, the expression levels of p-PI3K, p-Akt, and ABCA1 were decreased in THP-1 cell transferred with ABCA1 siRNA, but knockdown of miR-33-5p had an opposite effect. Furthermore, knockdown of miR-33-5p decreased the expression of MMP-2, MMP-9, TNF-α, total cellular cholesterol, and promoted cholesterol efflux in THP-1-derived foam cells. Importantly, LY294002 (PI3K inhibitor) or si-ABCA1 completely inhibited the stimulatory effects of miR-33-5p inhibitor. CONCLUSION: This study has found that knockdown of miR-33-5p induced ABCA1 expression and promoted inflammatory cytokines and cholesterol efflux likely via activating the PI3K/Akt signaling pathway.

Predictive Value of Serum Inflammatory Factors and FT3 for Stroke-Associated Pneumonia in Patients With Acute Ischemic Stroke.

OBJECTIVE: The ability of serum inflammatory factors and free triiodothyronine (FT3) in predicting the occurrence of stroke-associated pneumonia (SAP) in patients with acute ischemic stroke (AIS) was assessed in this study. METHODS: A retrospective analysis was conducted on 285 consecutive patients with AIS initially diagnosed and admitted to our hospital from January to December 2022. Patients were categorized into SAP and non-SAP groups based on the presence of SAP. Both groups were compared in terms of baseline characteristics, including National Institute of Health Stroke Scale (NIHSS) score, SAP risk assessment (A2DS2), TOAST classification. Independent risk factors for SAP were identified using multivariate logistic regression analysis, and the predictive value of inflammatory markers was evaluated through ROC curves. RESULTS: Among 285 patients with AIS, 40 (14.03%) were found to have developed SAP. Higher NIHSS and A2DS2 scores, elevated serum IL-1ß, IL-8, and IL-33 levels, increased age, atrial fibrillation, swallowing difficulties, and a higher proportion of patients with low FT3 levels were observed in the SAP group compared with the non-SAP group (all P<0.05). Significant risk factors for SAP in patients with AIS were identified through multivariate logistic regression analysis, including age, swallowing difficulties, NIHSS, A2DS2 , IL-1ß , IL-8 , IL-33, and FT3 (P<0.05). The highest predictive values were observed for A2DS2, FT3, and IL-8 with AUC values of 0.854, 0.844, and 0.823, respectively. CONCLUSION: SAP can be highly predicted by A2DS2, FT3, and IL-8, enabling the early identification of patients with high-risk SAP and facilitating timely intervention and treatment.

[Mechanism of large-conductance calcium-activated potassium channel involved in inflammatory response in sepsis].

OBJECTIVE: To explore the mechanisms of large-conductance calcium-activated potassium channel (BKCa) involved in inflammatory response in sepsis. METHODS: The serum levels of BKCa were measured by enzyme-linked immunosorbent assay (ELISA) in patients with sepsis (28 cases), patients with common infection (25 cases) and healthy people (25 cases). The relationship between levels of BKCa and acute physiology and chronic health evaluation II (APACHE II) were analyzed. Cultured RAW 264.7 cells were stimulated by lipopolysaccharide (LPS). In some experiments, a cell model of sepsis was constructed using Nigericin as the second stimulus signal. The mRNA and protein expressions of BKCa in RAW 264.7 cells stimulated with LPS (0, 50, 100, 1 000 µg/L) were measured by real-time fluorescence quantitative polymerase chain reaction (RT-qPCR) and Western blotting. RAW 264.7 cells were transfected with small interfering RNA of BKCa (siRNA-BKCa), and the levels of caspase-1 precursor (pro-caspase-1), interleukin-1ß precursor (pro-IL-1ß) in cell, and the levels of caspase-1 p20, IL-1ß p17 of cell culture medium, and NOD-like receptor protein 3 (NLRP3), nuclear factor-κB (NF-κB) were measured by Western blotting. The apoptosis were detected by staining with propidium iodide (PI), the release rate of lactate dehydrogenase (LDH) were measured, and the expression of apoptotic protein Gasdermin D (GSDMD) was measured by Western blotting to evaluate the effect of silencing BKCa on cell pyrosis. RESULTS: The level of serum BKCa in patients with sepsis was significantly higher than that in patients with common infection and health peoples (ng/L: 165.2±25.9 vs. 102.5±25.9, 98.8±20.0, both P < 0.05). In addition, the level of serum BKCa in patients with sepsis was significantly positively correlated with APACHE II score (r = 0.453, P = 0.013). LPS could construct a sepsis cell model by which LPS could promote BKCa expression in mRNA and protein with a concentration-dependent manner. The mRNA and protein expressions of BKCa in the cells stimulated by 1 000 µg/L LPS were significantly higher than that in the blank group (0 µg/L) [BKCa mRNA (2-ΔΔCt): 3.00±0.36 vs. 1.00±0.16, BKCa/ß-actin: 1.30±0.16 vs. 0.37±0.09, both P < 0.05]. Compared with the control group, the ratios of caspase-1 p20/pro-caspase-1 and IL-1ß p17/pro-IL-1ß in the model group were significantly increased (caspase-1 p20/pro-caspase-1: 0.83±0.12 vs. 0.27±0.05, IL-1ß p17/pro-IL-1ß: 0.77±0.12 vs. 0.23±0.12, both P < 0.05), however, transfection of siRNA-BKCa induced the decrease both of them (caspase-1 p20/pro-capase-1: 0.23±0.12 vs. 0.83±0.12, IL-1ß p17/pro-IL-1ß: 0.13±0.05 vs. 0.77±0.12, both P < 0.05). Compared with the control group, the number of apoptotic cells, LDH release rate and GSDMD expression in the model group were significantly increased [LDH release rate: (30.60±8.40)% vs. (15.20±7.10)%, GSDMD-N/GSDMD-FL: 2.10±0.16 vs. 1.00±0.16, both P < 0.05], however, transfection of siRNA-BKCa induced the decrease both of them [LDH release rate: (15.60±7.30)% vs. (30.60±8.40)%, GSDMD-N/GSDMD-FL: 1.13±0.17 vs. 2.10±0.16, both P < 0.05]. The mRNA and protein expressions of NLRP3 in sepsis cells were significantly higher than those in the control group [NLRP3 mRNA (2-ΔΔCt): 2.06±0.17 vs. 1.00±0.24, NLRP3/GAPDH: 0.46±0.05 vs. 0.15±0.04, both P < 0.05]. However, the expression of NLRP3 after siRNA-BKCa transfection was significantly lower than that in model group [NLRP3 mRNA (2-ΔΔCt): 1.57±0.09 vs. 2.06±0.17, NLRP3/GAPDH: 0.19±0.02 vs. 0.46±0.05, both P < 0.05]. Compared with the control group, the NF-κB p65 nuclear transfer of sepsis cell were significantly increased (NF-κB p65/Histone: 0.73±0.12 vs. 0.23±0.09, P < 0.05). However, the NF-κB p65 expression in the nucleus were decreased after siRNA-BKCa transfection (NF-κB p65/Histone: 0.20±0.03 vs. 0.73±0.12, P < 0.05). CONCLUSIONS: BKCa is involved in the pathogenesis of sepsis, and its possible mechanism is to activate NF-κB/NLRP3/caspase-1 signaling pathway to induce inflammatory factor production and cell death.

[A comparison of clinical characteristics between acute fatty liver of pregnancy and hemolysis, elevated liver enzymes and low platelets syndrome].

OBJECTIVE: To compare and analyze the clinical characteristics between acute fatty liver of pregnancy (AFLP) and the hemolysis, elevated liver enzymes and low platelets (HELLP) syndrome. METHODS: This is a retrospective cohort study. The clinical data of 13 cases with AFLP and 34 cases with HELLP syndrome were collected from three tertiary referral centers in Yunnan (the First Affiliated Hospital of Kunming Medical University, the Second Affiliated Hospital of Kunming Medical University, and Yan'an Hospital of Kunming City) from January 2016 to December 2021. The patients were diagnosed to AFLP and HELLP syndrome according to the Swansea criteria and the Tennessee classification system. The general characteristics, clinical features, laboratory results within 24 hours after admission, complications, maternal and neonatal outcomes were compared to analysis the differences between the two groups. RESULTS: (1) Maternal characteristics: compared with HELLP syndrome group, AFLP group had lower body mass index (BMI) and blood pressure at admission (both P < 0.01). (2) Clinical features: the most common symptoms in AFLP patients were skin jaundice, abdominal pain, nausea and vomiting, edema. The main manifestations of patients with HELLP syndrome were albuminuria, hypertension, edema, headache. Some patients had multiple symptoms concurrently. (3) Laboratory results: compared with HELLP syndrome group, the levels of platelet count (PLT), total bilirubin (TBil), direct bilirubin (DBil), γ-glutamyl transferase (γ-GGT), alkaline phosphatase (ALP), total bile acid (TBA), serum creatinine (SCr) and international standardized ratio (INR) in AFLP group were significantly increased within 24 hours after admission [PLT (×109/L): 107.69±51.13 vs.76.71±43.25,TBil (µmol/L): 121.60 (83.20, 170.00) vs.15.25 (7.22, 29.05), DBil (µmol/L): 86.50 (58.60, 104.00) vs. 4.30 (2.22,10.10), γ-GGT (U/L): 87.00 (37.00, 127.00) vs. 41.00 (19.00,64.42), ALP (U/L): 199.10 (109.00, 349.20) vs. 125.50 (90.50, 155.25), TBA (µmol/L): 51.50 (16.20, 117.40) vs. 4.15 (2.02, 6.95), SCr (µmol/L): 155.80 (129.00, 237.00) vs. 79.00 (65.43, 113.70), INR: 1.28 (1.17, 1.63) vs. 0.94 (0.88, 1.08), all P < 0.05], prothrombin time (PT) was significantly prolonged [seconds: 16.10 (14.50, 19.20) vs. 12.40 (11.43, 13.40), P < 0.05]. The level of blood glucose (GLU), fibrinogen (FIB) and the activity of antithrombin III (AT III) decreased significantly [GLU (mmol/L): 5.18±1.33 vs. 6.33±1.19, FIB (g/L): 1.96±1.46 vs. 3.81±1.58, AT III (%): 40.61±25.84 vs. 66.39±24.11, all P < 0.05]; (4) Complications: compared with HELLP syndrome group, the incidence of patients with hypoglycemia [30.77% (4/13) vs. 0% (0/34)], acute liver failure [53.85% (7/13) vs. 5.88% (2/34)], acute renal insufficiency [69.23% (9/13) vs. 8.82% (3/34)], coagulopathy [76.92% (10/13) vs. 38.24% (13/34)], disseminated intravascular coagulation (DIC) [53.85% (7/13) vs. 5.88% (2/34)], and multiple organ dysfunction syndrome (MODS) [53.85% (7/13) vs. 5.88% (2/34)] were significantly higher in AFLP group (all P <0.05). (5) Maternal and neonatal outcome: all patients delivered after admission. The total length of hospital and intensive care unit stay were significantly longer in the AFLP group than in the HELLP syndrome group [days: 17.00 (11.00, 25.00) vs. 9.00 (7.00, 12.00), 12.00 (4.00, 22.00) vs. 3.91 (0, 7.00), both P < 0.01]. Two AFLP patients died, including one due to intracranial venous thrombosis and one due to multiple organ failure and cardiopulmonary arrest. There were no deaths in the HELLP syndrome group. CONCLUSIONS: There are significant differences in maternal characteristics, laboratory results and complications between AFLP and HELLP syndrome. TBil, γ-GGT, SCr, FIB, INR and AT III activity may help to distinguish the two diseases.

[Clinical observation of extremely elevated erythrocyte sedimentation rate].

OBJECTIVE: To analyze the clinical features of adult patients with extremely elevated erythrocyte sedimentation rate (ESR, ESR ≥ 100 mm/1 h), so as improve the ability of clinicians to use erythrocyte sedimentation rate to assist in the diagnosis and treatment of diseases. METHODS: A retrospective cohort study was conducted to examine the clinical data of patients with ESR ≥ 100 mm/1 h admitted to the First Affiliated Hospital of Kunming Medical University from January 1st 2019 to December 31st 2019. The age, gender, clinical diagnosis, first ESR level after admission, blood routine, liver function, renal function, coagulation function and C-reactive protein (CRP) within 24 hours after admission were collected. Patient cohorts were divided into youth group (18-65 years old), middle-aged group (66-79 years old) and elderly group (≥ 80 years old) according to the new standards of human age classification of World Health Organization (WHO) 2019. Patient cohorts were also divided into infectious disease group, hematological disease group, autoimmune disease group, renal failure group and others according to their respective clinical diagnosis. The distribution of extremely elevated ESR in each group, and the correlation between ESR and various laboratory indicators were analyzed. RESULTS: (1) Among 429 patients with ESR ≥ 100 mm/1 h, there were 236 males and 193 females. There was no significant difference in ESR levels between males and females [mm/1 h: 108.00 (103.00, 119.75) vs. 117.00 (105.50, 140.00), P = 0.234]. (2) The age of 429 patients ranged from 18 to 98 years old, the average age was (53.70±18.70) years old. There were 310 cases in the youth group, 87 cases in the middle-aged group and 32 cases in the elderly group. The ESR level of the young group was significantly lower than that of the middle-aged group and the elderly group [mm/1 h: 108.00 (103.00, 120.00) vs. 119.00 (107.00, 140.00), 120.00 (110.25, 140.00), both P < 0.01]. (3) The main diagnoses associated with extremely elevated ESR were infectious diseases [157 cases (36.6%)], hematological system diseases [127 cases (29.6%)], autoimmune diseases [74 cases (17.2%)]. Pulmonary infection accounted for 58.0% (91/157) of infectious diseases. Hematopoietic stem cell diseases accounted for 45.7% (58/127), lymphocyte and plasma cell diseases accounted for [37.0% (47/127)] and erythrocyte diseases accounted for [11.0% (14/127)] of the hematological system diseases. Diffuse connective tissue diseases accounted for 75.7% (56/74) of autoimmune diseases. (4) Spearman correlation analysis showed that the extremely elevated ESR in all patients was significantly negatively correlated with the levels of red blood cell count (RBC), hemoglobin (HB) and hematocrit (HCT) (ρ value was -0.395, -0.381 and -0.383, respectively, all P < 0.01), the ESR was significantly positively correlated with the level of fibrinogen (FIB; ρ = 0.345, P < 0.01). A total of 266 patients were tested for both ESR and CRP, and there was no significantly correlation between ESR and CRP level (ρ = -0.019, P = 0.756). CONCLUSIONS: The extremely elevated ESR was more common in pulmonary infections diseases, hematopoietic stem cell diseases, lymphocyte and plasma cell diseases, erythrocyte diseases and diffuse connective tissue diseases. The extremely elevated ESR was significantly correlated with the levels of RBC, HB, HCT and FIB.

[The vicissitude of pathogenic bacteria isolated from critically ill patients in an intensive care unit during a period of using antibiotics].

OBJECTIVE: To observe the changing spectrum of the pathogenic bacteria during seven-day antibiotics targeted therapy in an intensive care unit (ICU). METHODS: In a group of 100 patients of hospital-acquired pneumonia (HAP) with identified pathogenic bacteria undergoing antibiotic treatment according to susceptibility test, the changes in the species of the pathogens and their ratio in their sputum specimens were studied, and the relationship were analyzed the characteristic between the changes and the age, the time of medication and the length of stay. RESULTS: Among all the bacterial isolates (n=295) in ICU, the percentage of Gram-negative bacillus was 62.4% (184/295). The prevalent causative microorganisms isolated were Pseudomonas aeruginosa 22.4% (66/295), MRSA/MRSE 20.7% (61/295) and Acinetobacter spp. 10.5% (31/295). When one or more than one potent antibiotic in accord with the result of sensitivity test, change in ratio of pathogens occurred in 160, and change in species in 126. When the use of antibiotics was prolonged, the change in the former became less often. The change in ratio was less in 3-5 days than that of 6-7 days, the ratio was 72.7%, 62.5%, 60.0% (P<0.01) respectively on the 3rd day, the 4th day and the 5th day, showing that susceptible pathogenic bacteria became less gradually, indicating that the treatment was effective . However, the change in species of pathogenic bacteria began more obvious, and it was more predominant on the 6th day and the 7th day, which was 66.0%, 77.1% (P<0.01) respectively, showing emergence of new non-susceptible pathogenic bacteria. With increase in the use of different antibiotics, the species of pathogenic bacteria showed to increase an increasing tendency of change. When Gram-negative bacillus infection was treated, antibiotic resistant bacteria such as Candida albicans and MRSA usually appeared. But when Gram-positive bacillus infections were treated, Candida albicans, Pseudomonas aeruginosa and Enterobacter cloacae readily appeared. There was relationship between the change in pathogenic bacteria and age, and the length of stay of the patients. The more older in age and the longer the length of stay, the change in pathogenic bacteria was more predominant. CONCLUSION: New antibiotic resistant pathogenic bacteria appears after seven-day antibiotic-targeted therapy in ICU. The change of species of pathogenic bacteria is related to the duration and type of using antibiotic, and also the age and length of stay. The longer time of use and the more different types of antibiotic used, the older in age and the longer in length of stay, the change in species of pathogenic bacteria is more predominant. Monitoring the dynamic change of pathogenic bacteria, adjusting the antibiotic promptly and rational use of antibiotics are very important to decrease the change in species and antibiotic resistance of the bacteria.

[Regulatorg Mchanism of MiR-152 on Proliferation, Metastasis and Tumorigenicity of SHI-1 Cells].

OBJECTIVE: To investigate the regulatory mechanism of miR-152 on proliferation, metastasis and tumorigenesis of human acute monocytic leukemia SHI-1 cells. METHODS: The purchased SHI-1 cell line was treated with miR-152 over-expression (miR-152 agomir group) or miR-152 inhibition (miR-152 antagomir group), and the negative control (NC) group was set up. The cell proliferation of each group was detected by CCK-8 assay. Scratch healing assay was employed to determine the migration of cells. Transwell assay was used to measure the invasion of cells. The expressions of Cyclin D1, Caspase-3, MMP-2, TIMP-2, E-cadherin and N-cadherin were detected by Western blot. The flow cyronetry with annexin V-FITC/PI double staining was applied to detect the cell apoptosis. The tumorigenesis of cells was examined by tumor formation experiment in nude mice. RESULTS: Compared with the NC group, the cell proliferation, migration and invasion ability in miR-152 agomir group were significantly decreased (P＜0.05), while that in miR-152 antagomir were significantly up-regulated (P＜0.05) . Compared with the NC group, the protein expression of Cyclin D1, MMP-2, N-cadherin were down-regulated in miR-152 agomir group, but the protein expression of Caspase-3, TIMP-2 and E-cadherin were all up-regulated siginificantly. At the same time, the apoptosis were enhanced, but the timorigenicity in nude mice were significantly decreased (all P＜0.05). The protein expression of Cyclin D1, MMP-2, N-cadherin in miR-152 antagomir group, showed high levels but Caspase-3, TIMP-2 and E-cadherin protein showed low levels in comparison with NC group. At the same time, the apoptosis was decreased but the timorigenicity in nude mice was significantly enhanced (all P＜0.05) . CONCLUSION: miR-152 can inhibit the proliferation, metastasis and tumorigenesis of SHI-1 cell line, at the same time induce cell apoptosis, thus providing a theoretical basis for the treatment of acute lymphoblastic leukemia.

Comparison between multi-slice spiral CT and magnetic resonance imaging in the diagnosis of peritoneal metastasis in primary ovarian carcinoma.

The advent of disease evaluation by means of multi-slice spiral computed tomography (MSCT) and magnetic resonance imaging (MRI) represents a continually emerging role in the evaluation of various diseases; however, its role is yet to be adequately defined. Thus, the aim of the study was to compare the diagnostic value of MSCT and MRI in the diagnosis of peritoneal metastasis in primary ovarian carcinoma. Between January 2013 and December 2015, MSCT or MRI data were collected from 42 patients who had been previously diagnosed with peritoneal metastasis of ovarian carcinoma at the First Affiliated Hospital of Kunming Medical University. The tumor location, size, edge, and shape were all evaluated independently by three qualified imaging physicians using a double-blind method to confirm whether the patients were indeed suffering from peritoneal metastasis, as well as to rank the metastatic lesions recorded on a five-point scale. It was hypothesized that MRI and MSCT were comparable in the evaluation of ovarian carcinoma. Therefore, a receiver operating characteristics (ROC) curve was used to analyze the results and also to directly compare the respective diagnostic values of MSCT and MRI. In total, 165 metastatic lesions were confirmed by means of surgical operation. MSCT revealed 131 metastatic lesions, while MRI confirmed 154 metastatic lesions. The metastatic sites were primarily located on the subphrenic, epiploon, and gastrocolic ligaments and were further confirmed by either MRI or CT. In regard to MSCT, the most common site of underdiagnoses was in the vicinity of the uterus-rectum-fossa. MRI displayed a high detection rate in every site. The omission diagnostic rate of MSCT and MRI were 20.61% and 6.67%, respectively, while the accuracy rates were 79.39% and 93.33%, respectively. The obtained results revealed that the MSCT value of area under the ROC curve was smaller than that for MRI. Our findings provided evidence asserting that MRI, in comparison to MSCT, was more accurate in diagnosing peritoneal metastasis in patients with ovarian carcinoma.