[Metformin suppresses hypoxia-inducible factor-1α expression in cancer-associated fibroblasts to block tumor-stromal cross-talk in breast cancer].

OBJECTIVE: To investigate the mechanism of metformin for regulating tumor-stromal cell cross-talk in breast cancer. METHODS: Tumor associated fibroblasts (CAFs) co-cultured with breast cancer cells were treated with metformin, and the changes in expressions of hypoxia-inducible factor-1α (HIF-1α), p-AMPK, stroma-derived factor-1 (SDF-1) and interleukin-8 (IL-8) in the CAFs were detected using ELISA, RT-qPCR or Western blotting; Transwell assay was used to evaluate the invasiveness of the tumor cells and its changes following treatment with exogenous SDF-1, IL-8 and TGF-ß1. The effects of HIF-1α shRNA or overexpression plasmid, AMPK shRNA, and treatment with OG (a proline hydroxylase inhibitor) or 2-OXO (a proline hydroxylase activator) were examined on p-AMPK, HIF-1α, SDF-1 and IL-8 expressions and invasiveness of the CAFs. RESULTS: Metformin treatment significantly increased the expression levels of p-AMPK, SDF-1 and IL-8 (P<0.05) and decreased HIF-1α expression (P<0.05) without affecting AMPK expression level (P>0.05) in the CAFs. The invasion ability of metformintreated breast cancer cells was significantly decreased (P<0.05). Exogenous SDF-1 and IL-8, HIF-1α overexpression, and OGinduced upregulation of HIF-1α all significantly attenuated the inhibitory effects of metformin on breast cancer cell invasion (P<0.05) and HIF-1α, SDF-1 and IL-8 expressions in CAFs (P<0.05). Transfection with HIF-1α shRNA or treatment with 2-OXO significantly decreased the invasiveness of breast cancer cells (P<0.05). P-AMPK knockdown significantly suppressed the inhibitory effect of metformin on HIF-1α expression in CAFs and on invasion of breast cancer cells (P<0.05). Treatment with TGF-ß1 partially decreased the inhibitory effect of metformin on HIF-1α expression in CAFs and invasiveness of the breast cancer cells (P<0.05). CONCLUSION: Metformin suppresses HIF-1α expression in CAFs to block tumor-stromal cross talk in breast cancer.

[Short-term substitution of calcineurin inhibitors (CNI) with recombinant humanized anti-CD25 monoclonal antibody (Basiliximab) as aGVHD prophylaxis in CNI intolerant patients after allogeneic hematopoietic stem cell transplantation].

Objectives: To investigate the efficacy of short-term substitution of recombinant humanized anti-CD25 monoclonal antibody (Basiliximab) as acute GVHD (aGVHD) prophylaxis in calcineurin inhibitors (CNI) intolerant patients following allogeneic hematopoietic stem cell transplantation (allo-HSCT) . Methods: This study included 17 patients with refractory malignant hematological disorders who underwent salvage allo-HSCT at the Bone Marrow Transplantation Department of Shanghai Zhaxin Traditional Chinese and Western Medicine Hospital from August 2021 to August 2022 and were treated with Baliximab to prevent aGVHD due to severe adverse reactions to CNI. There were seven men and ten women, with a median age of 43 years (18-67). Following the discontinuation of CNI, Basiliximab was administered at a dose of 1 mg/kg once weekly until CNI or mTOR inhibitors were resumed. Results: Basiliximab was started at an average of 5 (1-32) days after HSCT. The median duration of substitution was 20 (7-120) days. All had neutrophil engraftment within a median of 12 (10-17) days. Thirteen patients had platelet engraftment after a median of 13 (11-20) days. Four patients did not develop stable platelet engraftment. Eight patients (47.1% ) developed Grade Ⅱ-Ⅳ aGVHD, while four (23.6% ) developed Grade Ⅲ/Ⅳ aGVHD. Only one patient died from aGVHD. Before the end of the followup period, seven of 17 patients died. The longest followup period of the survivors was 347 days, and the median survival rate was not met. The overall survival (OS) rate at six months was 62.6%. Among the 17 patients, 13 (76.4% ) experienced cytomegalovirus reactivation, 7 (41.2% ) experienced EB virus activation, and no cytomegalovirus disease was observed. Conclusions: When CNI intolerance occurs during allo-HSCT, short-term replacement with Baliximab can be used as an alternative to prevent aGVHD.

Isolated non-immune mediated second-degree atrioventricular block in fetus: natural history and predictive factors for spontaneous recovery.

OBJECTIVES: To uncover the clinical course of fetal isolated non-immune mediated second-degree AVB and determine the factors associated with the spontaneous recovery for fetal non-immune second-degree atrioventricular block (AVB). METHODS: A total of 20 fetuses with isolated, non-immune mediated second-degree AVB were prospectively recruited between 2014 and 2022. These fetuses were divided into the spontaneous recovery group (n=12) and the non-spontaneous recovery group (n=8). Maternal and fetal basic characteristics, intrauterine and postnatal outcomes were compared between groups. RESULTS: Twelve fetuses restored 1:1 atrioventricular conduction in utero and did not recur during the postnatal follow-up period. The residual eight fetuses maintained as second-degree AVB and six of them were aborted due to parental request in utero. Of the two live children with second-degree AVB, one of them progressed to complete AVB at the latest follow up at the age of 34 months, but without any symptoms, heart enlargement or dysfunction. The residual one progressed to complete AVB and was finally diagnosed with type 2 long-QT syndrome. Fetuses in the spontaneous recovery group presented with earlier gestational age at diagnosis (20.0[17.0-26.0] vs. 24.5[18.0-35.0] weeks, p=0.004) and higher atrial rate (147[130-160] vs 138.00[125.00-149.00] bpm, p=0.006) in comparison with the non-spontaneous recovery group. A cut-off value of 22.5 weeks of gestational age and 144 bpm of atrial rate at diagnosis could predict the failure of spontaneous recovery, with sensitivities of 87.5%, 75%, and specificities of 92.0%, 87.5%, respectively. CONCLUSIONS: The outcome of fetal non-immune second-degree AVB was favorable. Earlier gestational age at diagnosis and higher atrial rate were related to spontaneous reversion for isolated non-immune-mediated second-degree AVB. However, prenatal gene test should be performed for those with persistent AVB to exclude the heritable disorders including LQTS. These findings may provide important references for clinical management and prenatal counseling. This article is protected by copyright. All rights reserved.

A Cholecystokinin Analogue Ameliorates Cognitive Deficits and Regulates Mitochondrial Dynamics via the AMPK/Drp1 Pathway in APP/PS1 Mice.

BACKGROUND: There are no drugs on the market that can reverse or slow Alzheimer's disease (AD) progression. A protease-resistant Cholecystokinin (CCK) analogue used in this study is based on the basic structure of CCK, which further increases the stability of the peptide fragment and prolongs its half-life in vivo. We observed a neuroprotective effect of CCK-8L in APPswe/PS1dE9 (APP/PS1) AD mice. However, its corresponding mechanisms still need to be elucidated. OBJECTIVE: This study examined CCK-8L's neuroprotective effects in enhancing cognitive impairment by regulating mitochondrial dynamics through AMPK/Drp1 pathway in the APP/PS1 AD mice. METHODS: Behavioural tests are applied to assess competence in cognitive functions. Transmission electron microscopy (TEM) was performed to observe the ultrastructure of mitochondria of hippocampal neurons, Immunofluorescent staining was employed to assay for Aß1-42, APP, Adenosine 5'-monophosphate (AMP)-activated protein kinase (AMPK) and dynamin-related protein1 (Drp1). CRISPR/Cas9 was utilized for targeted knockout of the CCKB receptor (CCKBR) in the mouse APP/PS1 hippocampal CA1 region. A model of lentiviral vector-mediated overexpression of APP in N2a cells was constructed. RESULTS: In vivo, experiments revealed that CCK analogue and liraglutide significantly alleviated cognitive deficits in APP/PS1 mice, reduced Aß1-42 expression, and ameliorated l damage, which is associated with CCKBR activation in the hippocampal CA1 region of mice. In vitro tests showed that CCK inhibited mitochondrial fission and promoted fusion through AMPK/Drp1 pathway. CONCLUSIONS: CCK analogue ameliorates cognitive deficits and regulates mitochondrial dynamics by activating the CCKB receptor and the AMPK/Drp1 pathway in AD mice.

[Analysis of the incidence and symptomatology of low anterior resection syndrome after laparoscopic anterior resection for rectal cancer].

Objective: This study aims to explore the temporal trend of Low Anterior Resection Syndrome (LARS) and its symptoms after laparoscopic anterior resection for rectal cancer. Methods: A retrospective cohort study design was employed. The study included primary rectal (adenocarcinoma) cancer patients who underwent laparoscopic anterior resection at Tongji Hospital, Huazhong University of Science and Technology, between January 1, 2010, and December 31, 2020. Complete medical records and follow-up data at 3, 6, 9, 12, and 18 months postoperatively were available for all patients. A total of 1454 patients were included, of whom 1094 (75.2%) were aged ≤65 years, and 597 (41.1%) were females. Among them, 1040 cases (71.5%) had an anastomosis-to-anus distance of 0-5cm, and 86 cases (5.9%) received neoadjuvant treatment. All patients completed the Chinese version of the LARS questionnaire and their LARS occurrence and specific symptom information were recorded at 3, 6, 9, 12, and 18 months postoperatively. Considering past literature and clinical experience, further subgroup analyses were performed to explore the potential impact factors on severe LARS, including anastomosis level, preoperative neoadjuvant therapy, postoperative adjuvant therapy, and the presence of preventive stoma. Results: The occurrence rates of LARS at 3, 6, 9, 12, and 18 months postoperatively were 78.5% (1142/1454), 71.4% (1038/1454), 55.0% (799/1454), 45.7% (664/1454), and 45.7% (664/1454), respectively (χ2=546.180, P<0.001). No statistically significant difference was observed between the 12-month and 18-month time points (P>0.05). When compared with the symptoms at 3 months, the occurrence rates of gas incontinence [1.7% (24/1454) vs. 33.9% (493/1454)], liquid stool incontinence [3.9% (56/1454) vs. 41.9% (609/1454)], increased stool frequency [79.6% (1158/1454) vs. 95.9% (1395/1454)], stool clustering [74.3% (1081/1454) vs. 92.9% (1351/1454)], and stool urgency [46.5% (676/1454) vs. 78.7% (1144/1454)] in the LARS symptom spectrum were significantly alleviated at 12 months (all P<0.05) and remained stable beyond 12 months (all P>0.05). With the extension of postoperative time, the incidence rates of severe LARS exhibited a decreasing trend in different subgroups, of anastomosis level, preoperative neoadjuvant therapy, postoperative adjuvant therapy, and the presence of preventive stoma, and reached stability at 12 months postoperatively (all P>0.05). Conclusion: LARS and its specific symptom profile showed a trend of gradual improvement over time up to 1 year postoperatively, and stabilized after more than 1 year. Increased stool frequency and stool clustering are the most common features of abnormal bowel dys function, which improve slowly after surgery.

The effect of post-traumatic long bone non-unions on health-related quality of life.

BACKGROUND: Fracture non-unions have a detrimental effect on patients due to reduced mobility and severe pain. Current literature on the quality of life in non-unions is limited, hence the purpose of this study, to quantify the health-related quality of life (HRQoL) among patients with post-traumatic long bone non-unions. It was hypothesized that the HRQoL of these patients is lower than the Dutch population standard as well as for multiple chronic diseases and musculoskeletal disorders. PATIENTS AND METHODS: From January 2020 to December 2021, this study included consecutive patients who were referred to a multidisciplinary (trauma, orthopedic and plastic surgery), non-union clinic at the Maastricht UMC+. All non-unions were evaluated using the Non-Union Scoring System (NUSS) questionnaire. Patient reported HRQoL outcomes were acquired using the EQ-5D-5L questionnaire and the Lower Extremity Functional Scale (LEFS). RESULTS: 50 patients were included, 18 females and 32 males, with a mean age of 55 years (± 15.5 SD). Eighteen patients presented with an open fracture, nine non-unions were infected and 39 patients had a lower extremity non-union. The mean NUSS score was 39.61 (± 14.6 SD). The mean EQ-5D-5L index score was 0.490 (± 0.261 SD), where patients experienced most problems with mobility. The mean EQ-5D-5L VAS was 61.4 (± 19.6 SD). The patients had a mean LEFS score of 28.7 (± 16.4 SD). The health-related quality of life was well below the age-corrected normative score of the Dutch population (EQ-5D-5L 0.857(p < 0.001); LEFS 77(p < 0.001)). This cohort's HRQoL was significantly lower than the HRQoL of multiple chronic and musculoskeletal disorders, including different forms of cancer and osteoarthritis. CONCLUSIONS: This study has quantified the detrimental effect of post-traumatic long bone non-unions on patient's health-related quality of life, being significantly lower than the HRQoL of the Dutch population as well as for multiple chronic and musculoskeletal medical conditions. This cohort demonstrates a patient population in need of more specialized care with a low health-related quality of life.

[Impact of immunosuppression intensified conditioning regimen for patients with strong positive pre-transplantation donor-specific anti-HLA antibodies (DSAs) undergoing haploidentical hematopoietic stem cell transplantation].

Objective: To explore the efficacy of immunosuppression intensified conditioning regimen in patients who have strongly positive donor-specific Anti-HLA antibodies (DSAs) and received a haploidentical hematopoietic stem cell transplantation (haplo-HSCT) . Methods: Clinical data of 10 patients with strongly positive pretransplant DSAs (defined as MFI ≥10000) were retrospectively analyzed in this study. All of them received a haplo-HSCT in the Hematology Department of Shanghai Zhaxin Traditional Chinese & Western Medicine Hospital. Results: ① Of all ten patients, three were males, and seven were females, with a median age of 53.5 (36-64) years. Of the 10 patients, three were diagnosed with acute myeloid leukemia, two were myelodysplastic syndromes (MDS), two were chronic myelomonocytic leukemia (CMML), two were in an accelerated phase of chronic myeloid leukemia (CML-AP), and one was primary myelofibrosis (PMF). ② Conditioning regimen consisted of fludarabine (Flu) /busulfan (Bu) combined with whole-body irradiation (TBI) /cyclophosphamide (Cy). ③ On the seventh day after transplantation, the median pretransplant DSA level was MFI 15 999 (10 210-23 417) and 10 787 (0-22 720). ④ Eight patients acquired hematopoietic reconstitution; the median time of neutrophil engraftment was 14 (10-16) days; and 18 (14-20) days for platelet engraftment. After a median follow-up of 12.5 (1.5-27) months, primary graft failure was found in one patient and another with poor graft function. Seven patients remained in a disease remission state, and all were DSA-negative. Conclusions: An intensified immunosuppression conditioning regimen can efficiently decrease the level of donor-specific anti-HLA antibodies (DSAs), leading to good short-term efficacy.

[Research progress on neurodevelopmental outcomes of small for gestational age infants].

The incidence of perinatal disease and perinatal mortality in small for gestational age infants increased significantly. This group of people is prone to a variety of long-term metabolic diseases and cardiovascular diseases, and is also prone to growth retardation and neurodevelopmental delay, which will seriously affect the long-term quality of life of children. The article studies the neurodevelopmental outcomes of small-for-gestational-age infants. By reviewing and sorting out previous literature, the neurodevelopmental disorders of small-for-gestational-age infants are analyzed according to five aspects: intellectual development, motor development, language development, sensory development, and mental illness. The classification and summary were carried out, and the influencing factors of neurodevelopmental disorders of SGA were also evaluated, so as to provide reference for promoting the improvement of neurodevelopmental outcomes of small-for-gestational-age infants.

[The role of methylation of placental glucocorticoid response gene in the association between pregnancy-related anxiety in the third trimester and birth outcomes].

Objective: To investigate the role of methylation of placental glucocorticoid response gene in the association between pregnancy-related anxiety in the third trimester and birth outcomes. Methods: Based on a prospective cohort study, singleton live births and their mothers from the Ma'anshan Birth Cohort Study (MABC) were included as participants in this study. The maternal pregnancy-related anxiety symptoms in the third trimester of pregnancy were evaluated by using the Pregnancy-related Anxiety Questionnaire. The neonatal birth outcomes were collected from medical records. The placental tissues from 300 pregnant women with pregnancy-related anxiety and 300 without pregnancy-related anxiety were collected to detect the methylation of FKBP5, NR3C1 and HSD11B2 genes using the Methyl Target approach. The methylation factors were extracted by exploratory factor analysis. Linear regression or logistic regression models were used to analyze the association between pregnancy-related anxiety in the third trimester, methylation factor scores, and birth outcomes. The mediating role of methylation factors in the association between pregnancy-related anxiety in the third trimester and birth outcomes was analyzed by using the Process procedure. Results: The mean age of 2 833 pregnant women was (26.60±3.60) years old. After adjusting for confounding factors, pregnancy-related anxiety in the third trimester increased the risk of small-for-gestational-age (OR=1.32, 95%CI:1.00-1.74). A total of 5 methylation factors were extracted, and the factor 5 was loaded with FKBP5 CpGs 18-21. Pregnancy-related anxiety in the third trimester was negatively correlated with the factor 5 (ß=-0.24,95%CI:-0.44--0.05). The factor 5 was positively correlated with the gestational age (ß=0.17, 95%CI:0.06-0.27). In addition, the factor 2 (ß=0.02,95%CI:0.00-0.04) and factor 3 (ß=0.03,95%CI:0.01-0.05) were positively correlated with 5-min Apgar score after delivery. However, this study did not found the mediating role of the scores of the factor characterized by FKBP5 in the relationship between pregnancy-related anxiety and birth outcomes. Conclusion: Pregnancy-related anxiety in the third trimester may reduce the methylation level of FKBP5 CpGs 18-21 in placental tissues and is associated with the risk of small-for-gestational-age.

Ncor1 Deficiency Promotes Osteoclastogenesis and Exacerbates Periodontitis.

Nuclear receptor corepressor 1 (Ncor1) has been reported to regulate different transcription factors in different biological processes, including metabolism, inflammation, and circadian rhythms. However, the role of Ncor1 in periodontitis has not been elucidated. The aims of the present study were to investigate the role of Ncor1 in experimental periodontitis and to explore the underlying mechanisms through an experimental periodontitis model in myeloid cell-specific Ncor1-deficient mice. Myeloid cell-specific Ncor1 knockout (MNKO) mice were generated, and experimental periodontitis induced by ligation using 5-0 silk sutures was established. Ncor1 flox/flox mice were used as littermate controls (LC). Histological staining and micro-computed tomography scanning were used to evaluate osteoclastogenesis and alveolar bone resorption. Flow cytometry was conducted to observe the effect of Ncor1 on myeloid cells. RNA sequencing was used to explore the differentially targeted genes in osteoclastogenesis in the absence of Ncor1. Coimmunoprecipitation (Co-IP), chromatin immunoprecipitation (ChIP) experiments, and dual luciferase assays were performed to explore the relationship between NCoR1 and the targeted gene. Alveolar bone resorption in the MNKO mice was significantly greater than that in the LC mice after periodontitis induction and osteoclastogenesis in vitro. The percentage of CD11b+ cells, particularly CD11b+ Ly6G+ neutrophils, was substantially higher in gingival tissues in the MNKO mice than in the LC mice. Results of RNA sequencing demonstrated that CCAAT enhancer binding protein α (Cebpα) was one of the most differentially expressed genes between the MNKO and LC groups. Mechanistically, Co-IP assays, ChIP experiments, and dual luciferase assays revealed that NCOR1 interacted with peroxisome proliferator-activated receptor gamma (PPARγ) and cooperated with HDAC3 to control the transcription of Cebpα. In conclusion, Ncor1 deficiency promoted osteoclast and neutrophil formation in mice with experimental periodontitis. It regulated the transcription of Cebpα via PPARγ to promote osteoclast differentiation.

[Interference of CTPS gene promotes toosendanin-induced apoptosis of human gastric cancer MKN-45 cells].

OBJECTIVE: To investigate the effect of interference of CTPS gene on toosendanin-induced apoptosis of gastric cancer MKN-45 cells. METHODS: Bioinformatic analysis was used to analyze CTPS gene expression in human gastric cancer tissues and the overall survival of gastric cancer patients with high CTPS gene expression. Human gastric cancer MKN-45 cells were transfected with a short hairpin interfering RNA targeting CTPS gene, and 48 h later, qRT-PCR and Western blotting were used to detect cellular expression CTPS at both the mRNA and protein levels. MKN-45 cells with CTPS knockdown were treated with 80 nmol/L toosendanin for 48 h, and the cell viability was assessed with MTT assay; the cell morphology was observed using laser confocal microscope, and the expression of γH2AX was detected with immunofluorescence assay. RESULTS: Bioinformatic analysis suggested that CTPS was highly expressed in human gastric cancer tissues, and gastric cancer patients with high CTPS gene expression had a shorter overall survival. MKN-45 cells transfected with Sh-CTPS interference vector showed significantly lowered cell survival rate (P < 0.01) with obvious cell shrinkage, irregular morphology, typical apoptotic changes, and increased cell apoptosis rate (P < 0.05). Treatment of the transfected cells with 80 nmol/L toosendanin for 48 h resulted in further reduction of the cell survival rate (P < 0.001), and the cells showed an increased apoptotic rate (P < 0.05) with appearance of apoptotic bodies. CONCLUSION: Interference of CTPS gene can promote TSN-induced apoptosis of gastric cancer MKN-45 cells.

[Detection and analysis of intestinal flora diversity in patients with complex anal fistula].

Objective: To explore the possibility that the intestinal flora profile in complex anal fistula patients is different to that of healthy controls. This was assessed by sequencing of 16S rDNA in fecal samples from cohorts representing these populations. Methods: Fecal samples were collected from 30 complex anal fistula patients and 30 matched healthy controls. Patients were included if they met the diagnostic criteria of cryptoglandular anal fistula and had exhibited symptoms for more than 3 months. Complex anal fistula is diagnosed under the following circumstances: if the fistula in question spans 2/3 or more of the diameter of the anal sphincter; if there are more than two external orifices or fistula tracks; or if recurrence is observed after previous anal fistula surgery. Patients were excluded if there were comorbities including inflammatory bowel disease (as assessed by colonoscopy), chronic diarrhea, chronic constipation, diabetes, gastrointestinal malignancies, liver/ kidney dysfunction, or cognitive impairment. Patients whose anal fistulas were caused by Crohn's disease, trauma, special infections (such as actinomycosis and tuberculosis) were also excluded, as were those who had used antibiotics, prebiotics, or probiotics that may affect intestinal microecology in the month prior to the study. Total bacterial genomic DNA was extracted by PCR amplification of the V4 hypervariable region of the 16S rRNA sequences. High-throughput sequencing and data analysis were performed on the Illumina Miseq platform. Finally, operational taxonomic unit (OTU) clustering, alpha diversity and LEfSE data analysis were carried out. The larger the Chao or ACE index is, the higher the species abundance of the microflora is expected to be. Similarly, a smaller value for the Simpson index or a larger value for the Shannon index indicates greater microflora diversity. There was no statistically significant difference in gender, age, body mass index (BMI), drinking history, or smoking history between the two groups (P>0.05), indicating that they were comparable. Results: The α-diversity analysis including ACE, Chao, Shannon and Simpson indexes indicated a richer diversity of intestinal microflora in complex anal fistula patients than in healthy controls. In both patients and controls, OUT cluster analysis demonstrated that 93.4%±32.0% and 87.4%±41.2% of sequences were from Firmicutes and Bacteroidetes spp., respectively. On a genus level, samples from anal fistula patients showed a greater abundance of Prevotella spp. (4.9%±7.4% vs. 0.1%±1.1%, P<0.001), Megamonas (3.9%±8.2% vs. 0.5%±4.2%, P<0.05) and Lachnospira (2.6%±5.7% vs. 0.1%±3.4%, P<0.05), while showing a lesser abundance of Proteobacteria spp. (0.02%±4.2% vs. 9.3%±14.4%, P<0.01), Enterococcus (0.02%±2.3% vs. 9.3%±19.6%, P<0.05), Bacteroides (24.7%±9.9% vs. 29.8%±9.1%, P<0.05) and Klebsiella (0.4%±4.2% vs. 3.9%±7.3%, P<0.05) compared with healthy controls. Intestinal flora diversity in the complex anal fistula group was richer than in controls, as indicated by a higher ACE index (293.30±44.00 vs. 218.75±33.83, t=102.069, P<0.001), a higher Chao index (318.40±41.99 vs. 250.00±46.38, t=77.818, P=0.028), a higher Shannon index (3.36±0.29 vs. 2.43±0.34, t=9.657, P=0.001), and a lower Simpson index (0.103±0.013 vs. 0.131±0.013, t=5.551, P=0.046). LDA effect size analysis suggests that the main strains of Veillonellaceae, Selenemondales and Negativicutes, which all belong to the phylum Firmicutes, have the greatest influence on the above difference (LDA>4). Conclusions: The diversity of intestinal flora in patients with complex anal fistula is greater than in healthy controls, suggesting that these bacteria or their metabolites may be involved in the occurrence and development of anal fistulas.

Ectopic thymic carcinoma of the parotid gland.

Ectopic thymic carcinoma (ETC) of the parotid gland is a rare entity. This report describes the case of a 52-year-old man with a painless mass in the right parotid gland. Magnetic resonance imaging (MRI) showed a biphasic mass consisting of a central segment and a peripheral segment. The patient underwent a superficial parotidectomy, and point-to-point correspondence sampling for analysis based on MRI findings was performed. The pathological finding was ETC, and there was an excellent association between MRI characteristics and histopathological findings. Subsequently, the patient underwent postoperative radiation therapy. At the 9-month follow-up, he had recovered well without facial paralysis, and there was no evidence of recurrence or metastasis. This report describes the clinical, radiological, and pathological features of the ETC.

Effectiveness of denosumab for fracture prevention in real-world postmenopausal women with osteoporosis: a retrospective cohort study.

To determine denosumab's effectiveness for fracture prevention among postmenopausal women with osteoporosis in East Asia, the risk of fracture was compared between patients continuing denosumab therapy versus patients discontinuing denosumab after one dose. The real-world effectiveness was observed to be consistent with the efficacy demonstrated in the phase III trial. INTRODUCTION: After therapeutic efficacy is demonstrated for subjects in global clinical trials, real-world evidence may provide complementary knowledge of therapeutic effectiveness in a heterogeneous mix of patients seen in clinical practice. This retrospective cohort study was conducted to compare the fracture risk in real-world clinical care received in Taiwan and Hong Kong between a treatment cohort (patients receiving denosumab 60 mg subcutaneously every 6 months) versus an off-treatment cohort (patients discontinuing after 1 dose of denosumab, which has no known clinical benefit) among real-world postmenopausal women. METHODS: This study included 38,906 and 2,835 postmenopausal women receiving denosumab in Taiwan and Hong Kong, respectively. The primary endpoint was hip fracture, and secondary endpoints were clinical vertebral and nonvertebral fractures. Propensity-score-matched analysis, adjusting for known covariates, was used to estimate hazard ratios (HRs) and 95% confidence intervals (CIs). The robustness of findings was evaluated with a series of sensitivity and quantitative bias analyses. RESULTS: In this study, 554 hip fractures were included in the primary Taiwan population analysis. The crude incidence rate was 0.9 per 100 person-years in the treatment cohort (n = 25,059) and 1.7 per 100 person-years in the off-treatment cohort (n = 13,847). After adjusting for prognostic differences between cohorts, denosumab reduced the risk of hip fractures by 38% (HR = 0.62, CI:0.52-0.75). Risk reductions of similar magnitude were observed for the secondary endpoints and for the analysis of the smaller Hong Kong population. CONCLUSION: The effectiveness of denosumab for fracture reduction among real-world postmenopausal women with osteoporosis was consistent with the efficacy demonstrated in a global clinical trial. REGISTRATION: EnCePP registration number: EUPAS26372; registration date: 12/11/2018.

Increased Agrobacterium-mediated transformation of Saccharomyces cerevisiae after deletion of the yeast ADA2 gene.

Agrobacterium tumefaciens is the causative agent of crown gall disease and is widely used as a vector to create transgenic plants. Under laboratory conditions, the yeast Saccharomyces cerevisiae and other yeasts and fungi can also be transformed, and Agrobacterium-mediated transformation (AMT) is now considered the method of choice for genetic transformation of many fungi. Unlike plants, in S. cerevisiae, T-DNA is integrated preferentially by homologous recombination and integration by non-homologous recombination is very inefficient. Here we report that upon deletion of ADA2, encoding a component of the ADA and SAGA transcriptional adaptor/histone acetyltransferase complexes, the efficiency of AMT significantly increased regardless of whether integration of T-DNA was mediated by homologous or non-homologous recombination. This correlates with an increase in double-strand DNA breaks, the putative entry sites for T-DNA, in the genome of the ada2Δ deletion mutant, as visualized by the number of Rad52-GFP foci. Our observations may be useful to enhance the transformation of species that are difficult to transform.

[Association between inflammation activity of left atrial epicardial adipose tissue measured by 18F-FDG PET/CT and atrial fibrillation].

Objective: To explore the association between inflammation activity of left atrial epicardial adipose tissue (LA-EAT) measured by 18F-fluorodeoxyglucose positron emission tomography/computed tomography (18F-FDG PET/CT) and atrial fibrillation (AF). Methods: A total of 78 patients with AF, who underwent 18F-FDG PET/CT in the Nuclear Medicine Department of the Third Affiliated Hospital of Soochow University due to abnormally elevated levels of tumor indicators or malignant tumors from March 2018 to December 2019, were enrolled in this retrospective study. According to the examination date of PET/CT and basic characteristics of AF patients (gender, age), a 1∶1 propensity score matching was used to enroll a non-AF control group (78 patients). The maximum standard uptake value of left atrial epicardial tissue (LA-EAT FDG SUVmax) and total EAT volume (V-EAT) were measured by 18F-FDG PET/CT. Left ventricular ejection fraction (LVEF) and left atrial diameter (LAD) were obtained by echocardiography. Blood lipids and biomarkers of inflammation were measured. The differences of clinical data and EAT-related indicators were compared between the AF group and control group. Logistic multivariate regression analysis was used to determine the related factors of AF. Then the receiver operating characteristic (ROC) curve was used to determine the cutoff value of LA-EAT FDG SUVmax on the diagnosis of AF. Univariate and multivariate logistic regression analysis were used to analyze the relationship between the increase of LA-EAT FDG SUVmax and AF. Results: The age was (66.9±10.2) years and there were 55 males (70.5%) in the AF group. The age was (66.9±8.0) years, and there were 52 males (66.7%) in the control group (both P>0.05). The LAD ((44.2±5.8) mm vs. (35.4±4.4) mm), V-EAT ((122.1±42.0) cm3 vs. (91.6±34.5) cm3), and LA-EAT FDG SUVmax ((1.6±0.3) vs. (1.4±0.2)) values were significantly higher, while LVEF ((60.1±4.7)% vs. (63.9±2.9)%) was lower in the AF group than in the control group (P all<0.001). Multivariate logistic regression analysis showed that LAD (OR=1.340, 95%CI 1.195-1.502), V-EAT (OR=1.016, 95%CI 1.001-1.031), and LA-EAT FDG SUVmax (OR=1.375, 95%CI 1.095-1.723) were positively correlated with AF, LVEF (OR=0.781, 95%CI 0.659-0.926) was negatively correlated with AF(P all<0.05). The area under the ROC curve of LA-EAT FDG SUVmax for diagnosis of AF was 0.680 (95%CI 0.597-0.764, P<0.001), and the best cut-off value was 1.415 with a sensitivity of 65.4% and specificity of 61.5%. After adjusting for high-density lipoprotein cholesterol, LVEF, LAD and V-EAT, LA-EAT FDG SUVmax≥1.415 was independently associated with AF (OR=2.982, 95%CI 1.122-7.926, P=0.010). Conclusions: The inflammatory activity of LA-EAT measured by 18F-FDG PET/CT is an independent risk factor of AF, and the increased inflammatory activity of LA-EAT is positively correlated with AF.

[Vascular endothelial growth factor promotes cancer stemness of triple-negative breast cancer via MAPK/ERK pathway].

OBJECTIVE: To investigate the role of vascular endothelial growth factor (VEGF) in regulating triple-negative breast cancer (TNBC) stem cells and the possible pathways involved in this regulatory mechanism. METHODS: The Oncomine database, UALCAN database and Human Protein Atlas (HPA) database were used to analyze the expression of VEGF in breast cancer and its association with the molecular subtypes and prognosis of breast cancer. Sphere formation assay was carried out to examine the effects of hVEGF165 on sphere formation ability of TNBC MDA-MB-231 cell line; Western blotting and RT-qPCR were performed to detect the expression of the tumor stem cell markers including CD44, c-Myc, Nanog, and ALDH1 and the activation of the related pathways. RESULTS: Data from the online databases all showed a significant increase of VEGF expression in breast cancer tissues than in the adjacent tissues (P < 0.0001), and its expression level was associated with the molecular subtypes of breast cancer. Specifically, the expression of VEGF was markedly higher in TNBC than in other subtypes of breast cancer. Survival analysis showed that breast cancer patients with a high VEGF expression had a significantly shortened overall survival (P < 0.0001). In the cell experiments, the sphere formation ability of MDA-MB-231 cells was significantly enhanced after treatment with hVEGF165 (P=0.0029). Compared with the monolayer cells, MDA-MB-231 spheres showed significantly increased expressions of VEGF, NRP-1, CD44, Nanog and c-Myc. Treatment with hVEGF165 resulted in significant time-dependent up-regulation of the expressions of CD44, c-Myc, Nanog and ALDH1 and down-regulation of CD24 expression in the cells. The results of Western blotting demonstrated that treatment with hVEGF165 caused significant activation of the ERK/MAPK pathway in MDA-MB-231 cells. CONCLUSION: VEGF promotes cancer stemness of triple-negative breast cancer possibly through the ERK/MAPK pathway.

[Changes of coagulation factor activity and its clinical significance in tumor patients with abnormal coagulation function].

Objective: To investigate the changes and clinical significance of coagulation factor activity in tumor patients with abnormal coagulation function. Methods: The clinical data of cancer patients who were hospitalized in Henan Cancer Hospital from January 2020 to June 2021 was collected. Thromboelastography (TEG) was used to monitor the coagulation function of tumor patients. Accordingly, 196 tumor patients with abnormal coagulation function were in the test group, and 36 tumor patients with normal status were in the control group. According to the coagulation index (CI) value of the TEG test results, the test group was divided into two groups: hypercoagulability test group (n=104): CI value>3; hypocoagulation test group (n=92): CI value<-3. Meanwhile, each test group was divided into 3 subgroups according to the R value, K value, α angle and MA value of the TEG results, namely hypercoagulable group one (n=37), hypercoagulable group two (n=34), and hypercoagulable group three (n=33); hypocoagulation group one (n=33), hypocoagulation group two (n=30), and hypocoagulation group three (n=29). The activities of coagulation factors (F) Ⅱ, Ⅴ, Ⅶ, Ⅷ, Ⅸ, Ⅹ, Ⅺ, Ⅻ and von willebrand factor (vWF) were measured and compared for all patients in the test groups and the control group in the same period. Results: Compared with the normal control group, the hypercoagulable group one showed higher FⅡ, FⅤ, FⅦ, FⅧ, FⅪ and vWF activity, and the values were (1 105±281), (1 352±326), (1 628±397), (1 795±314), (1 389±288) and (1 908±486) U/L, respectively (P<0.01). The hypercoagulable group two showed higher FⅡ, FⅤ, FⅦ, FⅧ FⅩ and vWF activity, and the values were (1 068±189), (1 194±205), (1 529±394), (1 562±241), (1 150±196) and (1 722±415) U/L, respectively (P<0.05). Hypercoagulable group three showed higher FⅦ, FⅩ and vWF activity, and the values were (1 411±196), (1 212±327) and (1 713±457) U/L, respectively (P<0.01). In contrast, the hypocoagulation group one showed lower FⅤ, FⅧ, FⅨ, FⅫ and vWF activity, and the values were (732±96), (695±64), (1 216±191), (832±128) and (1 088±117) U/L, respectively (P<0.05). Hypocoagulation group two showed lower FⅤ, FⅦ, FⅧ and FⅫ activity, and the values were (714±102), (1 125±108), (783±95) and (912±111) U/L, respectively (P<0.01). Hypocoagulation group three had lower FⅪ, FⅫ and vWF activity, and the values were (812±92), (827±179) and (916±76) U/L, respectively (P<0.01). Conclusions: Only part of the coagulation factor activity changes significantly in the tumor patients with abnormal coagulation function. In tumor patients with hypercoagulable state, the high activity of FⅡ and FⅩ becomes an important factor in anticoagulant therapy, while high FⅤ, FⅧ activity can cause deep vein thrombosis. In the hypocoagulation state, the significant decreases of FⅤ and FⅧ activity often cause bleeding or oozing.