Three-dimensional chromatin analysis reveals Sp1 as a mediator to program and reprogram HPV-host epigenetic architecture in cervical cancer.

Human papillomavirus (HPV) is predominantly associated with HPV-related cancers, however, the precise mechanisms underlying the HPV-host epigenetic architectures in HPV carcinogenesis remain elusive. Here, we employed high-throughput chromosome conformation capture (Hi-C) to comprehensively map HPV16/18-host chromatin interactions. Our study identified the transcription factor Sp1 as a pivotal mediator in programming HPV-host interactions. By targeting Sp1, the active histone modifications (H3K27ac, H3K4me1, and H3K4me3) and the HPV-host chromatin interactions are reprogrammed, which leads to the downregulation of oncogenes located near the integration sites in both HPV (E6/E7) and the host genome (KLF5/MYC). Additionally, Sp1 inhibition led to the upregulation of immune checkpoint genes by reprogramming histone modifications in host cells. Notably, humanized patient-derived xenograft (PDX-HuHSC-NSG) models demonstrated that Sp1 inhibition promoted anti-PD-1 immunotherapy via remodeling the tumor immune microenvironment in cervical cancer. Moreover, single-cell transcriptomic analysis validated the enrichment of transcription factor Sp1 in epithelial cells of cervical cancer. In summary, our findings elucidate Sp1 as a key mediator involved in the programming and reprogramming of HPV-host epigenetic architecture. Inhibiting Sp1 with plicamycin may represent a promising therapeutic option for HPV-related carcinoma.

Specific patterns and potential risk factors to predict 3-year risk of death among non-cancer patients with advanced chronic kidney disease by machine learning.

Chronic kidney disease (CKD) is a major public health concern. But there are limited machine learning studies on non-cancer patients with advanced CKD, and the results of machine learning studies on cancer patients with CKD may not apply directly on non-cancer patients. We aimed to conduct a comprehensive investigation of risk factors for a 3-year risk of death among non-cancer advanced CKD patients with an estimated glomerular filtration rate < 60.0 mL/min/1.73m2 by several machine learning algorithms. In this retrospective cohort study, we collected data from in-hospital and emergency care patients from 2 hospitals in Taiwan from 2009 to 2019, including their international classification of disease at admission and laboratory data from the hospital's electronic medical records (EMRs). Several machine learning algorithms were used to analyze the potential impact and degree of influence of each factor on mortality and survival. Data from 2 hospitals in northern Taiwan were collected with 6565 enrolled patients. After data cleaning, 26 risk factors and approximately 3887 advanced CKD patients from Shuang Ho Hospital were used as the training set. The validation set contained 2299 patients from Taipei Medical University Hospital. Predictive variables, such as albumin, PT-INR, and age, were the top 3 significant risk factors with paramount influence on mortality prediction. In the receiver operating characteristic curve, the random forest had the highest values for accuracy above 0.80. MLP, and Adaboost had better performance on sensitivity and F1-score compared to other methods. Additionally, SVM with linear kernel function had the highest specificity of 0.9983, while its sensitivity and F1-score were poor. Logistic regression had the best performance, with an area under the curve of 0.8527. Evaluating Taiwanese advanced CKD patients' EMRs could provide physicians with a good approximation of the patients' 3-year risk of death by machine learning algorithms.

RNA-binding protein RPS7 promotes hepatocellular carcinoma progression via LOXL2-dependent activation of ITGB1/FAK/SRC signaling.

BACKGROUND: Metastasis is one of the leading cause contributes to treatment failure and poor prognosis of hepatocellular carcinoma (HCC) patients. The underlying mechanism of HCC metastasis remains to be determined. Although several RNA binding proteins (RBPs) have been found to participate in tumorigenesis and progression of liver cancer, the role of RBPs in HCC patients with extrahepatic metastases is poorly understood. METHODS: By performing RNA-seq of primary HCC tissues (including HCC with extrahepatic metastasis and those did not develop metastasis), we identified a set of HCC metastasis-associated RBPs candidates. Among which, ribosomal protein S7 (RPS7) was found to be remarkably increased in HCC tissues and be strongly related to HCC poor survival. Overexpression or CRISPR-Cas9-mediated knockout were applied to investigate the role of RPS7 on the metastasis-associated phenotypes of HCC cells. RNA sequencing, RIP, RNA-pull down, dual luciferase reporter assay, nascent RNA capture assay, and RNA decay and so on, were applied to reveal the underlying mechanism of RPS7 induced HCC metastasis. RESULTS: Gain- and loss- of function analyses revealed that RPS7 promoted HCC cells adhesion, migration and invasion capabilities, as well as lung metastasis. Mechanistically, we uncovered that lysyl oxidase-like 2 (LOXL2) was a critical downstream target of RPS7. RPS7 could stabilize LOXL2 mRNA by binding to AUUUA motifs in the 3155-3375 region of the 3'UTR of LOXL2 mRNA, thus increased LOXL2 expression via elevating LOXL2 mRNA abundance. Further research revealed that LOXL2 could accelerate focal adhesion formation through maintaining the protein stability of ITGB1 and activating ITGB1-mediated FAK/SRC signaling pathway, and thereby contribute to the pro-metastasis effect of RPS7. CONCLUSIONS: Taken together, our data reveal a novel function of RPS7 in HCC metastasis, also reveal the critical roles of the RPS7/LOXL2/ITGB1 axis in HCC metastasis and shed new light on the exploration of molecular drugs against HCC.

Ex Vivo Evaluation of Combination Immunotherapy Using Tumor-Microenvironment-on-Chip.

Combination immunotherapy has emerged as a promising strategy to address the challenges associated with immune checkpoint inhibitor (ICI) therapy in breast cancer. The efficacy of combination immunotherapy hinges upon the intricate and dynamic nature of the tumor microenvironment (TME), characterized by cellular heterogeneity and molecular gradients. However, current methodologies for drug screening often fail to accurately replicate these complex conditions, resulting in limited predictive capacity for treatment outcomes. Here, a tumor-microenvironment-on-chip (TMoC), integrating a circulation system and ex vivo tissue culture with physiological oxygen and nutrient gradients, is described. This platform enables spatial infiltration of cytotoxic CD8+ T cells and their targeted attack on the tumor, while preserving the high complexity and heterogeneity of the TME. The TMoC is employed to assess the synergistic effect of five targeted therapy drugs and five chemotherapy drugs in combination with immunotherapy, demonstrating strong concordance between chip and animal model responses. The TMoC holds significant potential for advancing drug development and guiding clinical decision-making, as it offers valuable insights into the complex dynamics of the TME.

Phytotoxic and Antimicrobial Terrein Derivatives and Butenolides Isolated from the Endophytic Fungus Aspergillus terreus HT5.

Two new terrein derivatives, aspergilethers A and B (1 and 2), two known analogues (3 and 4), and three known butenolides (5-7) were isolated from the endophyte Aspergillus terreus HT5. Their structures were determined by spectroscopic analysis and ECD and NMR calculations. Interestingly, 1 and 2 had unpresented medium aliphatic side chains in terrein derivatives, with different absolute configurations at C-7, which was very scarce. (+)-Terrein (3) exhibited potent postemergence phytotoxicity toward Amaranthaceae, Portulacaceae, and Fabaceae, with MIC values of 250-1000 µg/mL. Transcriptome analysis and qRT-PCR suggested that (+)-terrein induced the transcriptional expression of aging-related genes to accelerate organ senescence and stimulated plant detoxification response. The conjugated system between keto carbonyl and double bonds in the cyclopentenone ring and side chain, and the configurations of C-2 and C-3, played critical roles in the phytotoxicity of terrein derivatives. Meanwhile, 3 was first reported to display moderate antioomycetes activity toward Phytophthora nicotiana.

Single-cell RNA sequencing reveals the heterogeneity and intercellular communication of hepatic stellate cells and macrophages during liver fibrosis.

Uncontrolled and excessive progression of liver fibrosis is thought to be the prevalent pathophysiological cause of liver cirrhosis and hepatocellular cancer, and there are currently no effective antifibrotic therapeutic options available. Intercellular communication and cellular heterogeneity in the liver are involved in the progression of liver fibrosis, but the exact nature of the cellular phenotypic changes and patterns of interregulatory remain unclear. Here, we performed single-cell RNA sequencing on nonparenchymal cells (NPCs) isolated from normal and fibrotic mouse livers. We identified eight main types of cells, including endothelial cells, hepatocytes, dendritic cells, B cells, natural killer/T (NK/T) cells, hepatic stellate cells (HSCs), cholangiocytes and macrophages, and revealed that macrophages and HSCs exhibit the most variance in transcriptional profile. Further analyses of HSCs and macrophage subpopulations and ligand-receptor interaction revealed a high heterogeneity characterization and tightly interregulated network of these two groups of cells in liver fibrosis. Finally, we uncovered a profibrotic Thbs1+ macrophage subcluster, which expands in mouse and human fibrotic livers, activating HSCs via PI3K/AKT/mTOR signaling pathway. Our findings decode unanticipated insights into the heterogeneity of HSCs and macrophages and their intercellular crosstalk at a single-cell level, and may provide potential therapeutic strategies in liver fibrosis.

Matching intraoperative teaching and learning for medical undergraduates via modified briefing-intraoperative teaching-debriefing (BID) model.

Intraoperative teaching is a challenging task. The briefing-intraoperative teaching-debriefing (BID) model, which is based on guided discovery learning at limited time intervals, has rarely been investigated. This study validated the benefits of the modified BID model on medical clerks. This study involved 37 first-year medical clerks enrolled from September 2019 to May 2020. Every learner scrubbed in one the totally implantable venous access device placement surgery and completed a pre-/posttest survey on surgical procedures and associated anatomy conducted through an intraoperative teaching questionnaire. Of these participants, 15 merely observed throughout the entire procedure (observation group), whereas the remaining 22 performed simple suturing under supervision (suturing group). All participants underwent an objective structured assessment of simple interrupted suturing skills at the end of the observership. Correlations were tested using a two-tailed paired t-test, with a p-value < 0.05 indicating statistical significance. The response rate was 100% and participants could reconfirm the precise venous access, catheter tip location, and suture materials for portal fixation after totally implantable venous access device placement (p < 0.05). Although a relatively higher satisfaction of the intraoperative teaching environment and educator attitude was reported in the suturing group than in the observation group, the difference in scores on the objective structured assessment was not statistically significant (8.7 ± 1.8 vs. 7.2 ± 3.7; p = 0.104). Our findings indicate that the modified BID model with hands-on experience is a practicable module for matching intraoperative teaching and learning via learning perception enhancement for medical undergraduates during totally implantable venous access device placement.

Network meta-analysis of first-line immune checkpoint inhibitor therapy in advanced non-squamous non-small cell lung cancer patients with PD-L1 expression ≥ 50.

INTRODUCTION: The optimal first-line immunotherapy regimen for advanced non-squamous non-small cell lung cancer (NS-NSCLC) patients with programmed cell death ligand 1 (PD-L1) expression ≥ 50% remains unclear. Our aim is to determine the most effective treatment regimen through a network meta-analysis (NMA) comparing these treatments. METHODS: A systematic search was performed in PubMed, Cochrane Library, Web of Science, and Embase databases, and a Bayesian network meta-analysis was conducted. To ensure transparency, the study was registered in the International Prospective Register of Systematic Reviews (CRD42022349712). RESULTS: The analysis included 11 randomized controlled trials (RCTs) with 2037 patients and 12 immunotherapy combinations. ICI-ICI, ICI alone, and chemotherapy-ICI showed significant advantages over chemotherapy in terms of overall survival (OS) and progression-free survival (PFS). Pembrolizumab plus chemotherapy showed the best OS results compared to chemotherapy. Tislelizumab plus chemotherapy and sintilimab plus chemotherapy provided the best PFS results. CONCLUSIONS: For NS-NSCLC patients with PD-L1 ≥ 50%, pembrolizumab plus chemotherapy, tislelizumab plus chemotherapy, and sintilimab plus chemotherapy are recommended as good treatment options based on the results of this Network meta-analysis (NMA).

The Effects of Angiosome Morphology on Choke Vessels and Flap Necrosis in a Rat Multiterritory Perforator Flap.

BACKGROUND: Although the angiosome concept has been proposed for a long time, very few studies have been done on its morphology. Our study investigated the effects of angiosome morphology on choke vessels and flap necrosis in a rat multiterritory perforator flap. METHODS: Seventy-two male Sprague-Dawley rats were randomly divided into 3 groups (n = 24/group). The flap contained the right iliolumbar, posterior intercostal, and thoracodorsal angiosomes (TDAVs), termed angiosomes I, II, and III, respectively. Only the posterior intercostal artery and iliolumbar vein were preserved in group 1, whereas only the posterior intercostal artery and vein were preserved in group 2, and only the posterior intercostal artery and thoracodorsal vein were preserved in group 3. Distances from angiosome II to angiosome I (II-I), angiosome II to angiosome III (II-III), angiosome I to the caudal side of the flap (I-caudal), and angiosome III to the cranial side of the flap (III-cranial) were measured. Arteriography, flap necrosis, average microvascular density, and vascular endothelial growth factor expression were evaluated. RESULTS: The II-I distance was significantly greater than that of II-III (3.853 ± 0.488 versus 3.274 ± 0.433 cm, P = 0.012), whereas the distance of I-caudal resembled that of III-cranial (1.062 ± 0.237 versus 0.979 ± 0.236 cm, P = 0.442). The iliolumbar and posterior intercostal angiosomes were multidirectional, whereas the TDAV was craniocaudal and unidirectional. Seven days after the operation, the choke arteries had transformed into true anastomotic arteries. Flap necrosis was lowest in group 3, followed by group 2, and highest in group 1 (10.5% ± 2.4% versus 18.3% ± 3.5% versus 25.5% ± 4.6%, P < 0.01), whereas group 3 showed the highest microvascular density and vascular endothelial growth factor expression, in contrast to groups 2 and 1, with the lowest. CONCLUSIONS: The choke vessel adjacent to the craniocaudal and unidirectional TDAV significantly blocked venous return. Increasing venous return may reduce the necrosis.

High-pressure infusion improves multi-territory perforator flap viability via choke artery dilation: A preliminary study in a rat model.

BACKGROUND: Multi-territory perforator flaps have become the preferred option for the repair and reconstruction of large soft tissue defects. Although methods (e.g., pharmacological agents, mechanical stimulation, and thermal stimulation) were developed to open choke vessels to improve flap survival, the flap necrosis rate is still as high as 28.8%. The authors hypothesized that high-pressure infusion might enhance flap viability by dilating choke arteries intraoperatively in a rat model of multi-territory perforator flap. METHODS: Two-month-old male Sprague-Dawley rats were randomized into two groups (n = 32 each). During the multi-territory perforator flap elevation based on the right superficial epigastric angiosome, one group received continuous high-pressure infusion (mean pressure, 250 mmHg; duration, 1 min) of an isotonic heparin sodium solution (12,500 U/L) via the artery in the pedicle, whereas the other group received no infusion. At 7 days postoperatively, arteriography was performed; endothelial nitric oxide synthase (eNOS) and vascular endothelial growth factor (VEGF) expression and microvascular density were evaluated by western blot and histology, respectively; and flap survival was compared. Moreover, intraluminal diameters were examined at 1 day and 7 days postoperatively using hematoxylin and eosin staining, and coagulation function was assessed immediately postoperatively. RESULTS: High-pressure infusion significantly promoted the dilation of choke arteries at 1 day and 7 days postoperatively. It also increased eNOS and VEGF expression, flap survival, and microvascular density. The coagulation function remained unaffected. CONCLUSIONS: High-pressure infusion allowed intraoperative and postoperative dilation of the choke arteries that enhanced the viability of multi-territory perforator flaps in rats.

TMEM147 Correlates with Immune Infiltration and Serve as a Potential Prognostic Biomarker in Hepatocellular Carcinoma.

Background: Hepatocellular carcinoma (HCC) is one of the most prevalent malignancies and is associated with high mortality. Transmembrane protein 147 (TMEM147) is a seven-transmembrane protein that may mediate immune regulation. However, the relevance of TMEM147 to immune regulation in HCC and the prognosis of HCC patients are unclear. Methods: We analyzed TMEM147 expression in HCC by using the Wilcoxon rank-sum test. Real time quantitative PCR (RT-qPCR) and Western blot analysis of tumor tissues and cell lines were used to verify TMEM147 expression in HCC. The influence of TMEM147 on HCC prognosis was assessed using Kaplan-Meier analysis, Cox regression analysis, and a prognostic nomogram. The functions of the TMEM147-related differentially expressed genes (DEGs) were identified by Gene Ontology (GO)/Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses and gene set enrichment analysis (GSEA). In addition, we examined the associations between TMEM147 expression and immune infiltration using single-sample gene set enrichment analysis (ssGSEA) and immunofluorescence staining of HCC tissues. Results: Our results showed that the expression of TMEM147 was significantly higher in human HCC tissues than in adjacent normal liver tissues, with similar findings in human HCC cell lines. High TMEM147 expression was correlated with T stage, pathological stage, histological grade, race, alpha-fetoprotein level, and vascular invasion in HCC. Moreover, we revealed that high TMEM147 expression was associated with shorter survival times and that TMEM147 could be a risk factor for overall survival, along with T stage, M stage, pathological stage, and tumor status. Mechanistic studies revealed that high TMEM147 expression was linked to the B lymphocyte, antigen response, IL6 signaling pathway, cell cycle, Kirsten rat sarcoma viral oncogene homolog (KRAS) signaling pathway, and myelocytomatosis oncogene (MYC) targets. Correspondingly, TMEM147 expression was positively associated with the infiltration of immune cells, including Th2 cells, follicular helper T cells, macrophages, and NK CD56 bright cells in HCC. Conclusions: TMEM147 might be a biomarker for poor prognosis and is related to immune cell infiltration in HCC.

Anlotinib Combined With Toripalimab as First-Line Therapy for Unresectable Hepatocellular Carcinoma: A Prospective, Multicenter, Phase II Study.

BACKGROUND: For patients with unresectable hepatocellular carcinoma (HCC), the first-line therapeutic options are still relatively limited, and treatment outcomes remain poor. We aimed to assess the efficacy and safety of anlotinib combined with toripalimab as first-line therapy for unresectable HCC. METHODS: In this single-arm, multicenter, phase II study (ALTER-H-003), patients with advanced HCC without previous systemic anticancer therapy were recruited. Eligible patients were given anlotinib (12 mg on days 1-14) combined with toripalimab (240 mg on day 1) in a 3-week cycle. The primary endpoint was the objective response rate (ORR) by immune-related Response Evaluation Criteria in Solid Tumours (irRECIST)/RECIST v1.1 and modified RECIST (mRECIST). Secondary endpoints included disease control rate (DCR), duration of response (DoR), progression-free survival (PFS), overall survival (OS), and safety. RESULTS: Between January 2020 and Jul 2021, 31 eligible patients were treated and included in the full analysis set. At data cutoff (January 10, 2023), the ORR was 29.0% (95% CI: 12.1%-46.0%) by irRECIST/RECIST v1.1, and 32.3% (95% CI: 14.8%-49.7%) by mRECIST criteria, respectively. Confirmed DCR and median DoR by irRECIST/RECIST v1.1 and mRECIST criteria were 77.4 % (95% CI: 61.8%-93.0%) and not reached (range: 3.0-22.5+ months), respectively. Median PFS was 11.0 months (95% CI: 3.4-18.5 months) and median OS was 18.2 months (95% CI: 15.8-20.5 months). Of the 31 patients assessed for adverse events (AEs), the most common grade ≥ 3 treatment-related AEs were hand-foot syndrome (9.7%, 3/31), hypertension (9.7%, 3/31), arthralgia (9.7%, 3/31), abnormal liver function (6.5%, 2/31), and decreased neutrophil counts (6.5%, 2/31). CONCLUSIONS: Anlotinib combined with toripalimab showed promising efficacy and manageable safety in Chinese patients with unresectable HCC in the first-line setting. This combination therapy may offer a potential new therapeutic approach for patients with unresectable HCC.

Bacterial diversity of middle ear cholesteatoma by 16S rRNA gene sequencing in China.

In this study, the bacterial diversity of acquired middle ear cholesteatoma (MEC) was evaluated to reveal its pathogenesis and provides a guide for the use of antibiotics. Twenty-nine cases of acquired MEC and eight cases of healthy middle ears undergoing cochlear implantation (CI) were evaluated. Full-length 16S rRNA gene sequencing was performed to profile the bacterial communities in lesions and healthy tissues of the middle ear. ACE (P = 0.043) and Chao1 (P = 0.039) indices showed significant differences in alpha diversity (P < 0.05). Analysis of PERMANOVA/Anosim using the Bray-Curtis distance matrix results suggested that the between-group differences were greater than the within-group differences (R = 0.238, P < 0.05, R2 = 0.066, P < 0.05). Bacterial community analysis revealed that Alphaproteobacteria at the class level and Caulobacterales and Sphingomonadales at the order level were significantly different (P < 0.05). In the LefSe (Linear discriminant analysis effect size) analysis, Porphyromonas bennonis was elevated, and Bryum argenteum and unclassified Cyanobacteriales were reduced at the species level in MEC (P < 0.05). Fifteen metabolic pathways were found to be significantly different between the two groups by analysing the abundance of metabolic pathways in level 2 of the Kyoto Encyclopaedia of Genes and Genomes (KEGG). Seven and eight metabolic pathways were significantly elevated in the MEC and control groups, respectively (P < 0.05). The role of bacteria in the pathogenesis of acquired MEC was further refined through analysis of metabolic pathways. These findings indicate that the acquired MEC and healthy middle ear contain more diverse microbial communities than previously thought.

Efficacy of a fingertip reconstruction technique using the tongue-shaped flap and advancement flap for repairing congenital syndactyly with osseous fusion of the distal phalanges.

BACKGROUND: To explore the clinical efficacy of using tongue-shaped flaps and advancement flaps to reconstruct the fingertips in congenital syndactyly patients with osseous fusion of the distal phalanges. METHODS: From January 2016 to January 2019, 12 patients with congenital syndactyly, involving 30 digits in total, presented to our hospital and were surgically treated with tongue-shaped flaps, as well as with advancement flaps to reconstruct the fingertips. The flap infection rate, necrosis rate and any other early complications were recorded. Fingertip aesthetics were reported according to the modified Bulic scale. A questionnaire was used to assess the satisfaction of the patients' family members. RESULTS: All cases were thoroughly reviewed. The postoperative period for inclusion in this study ranged from 36 to 60 months, with an average follow-up time of 45 months. During this period, no complications such as infection and/or necrosis of any flap were observed. Significant improvements in finger aesthetics and functioning compared to preoperative values were observed in most cases. Based on the modified Bulic scale, of 30 fingertips, an excellent result was obtained for 3, a very good result for 13, a good result for 13 and a poor result for just 1. Family members were satisfied with the treatment outcome. CONCLUSIONS: This technique employing tongue-shaped flaps and advancement flaps to reconstruct fingertips is effective, which enables the attainment of favourable aesthetic and functional outcomes in congenital syndactyly patients with osseous fusion of the distal phalanges.

Immune-Related Colitis Induced by Camrelizumab: A Case Report.

In recent years, immunotherapy has become a major research focus in the field of cancer treatment. Because of its good efficacy and lasting immune response, immune checkpoint inhibitors have benefited the long-term survival of many types of cancer patients. However, overactivation of the immune system may attack normal organs and cause a series of immune related adverse reactions. Among them, due to the high incidence of immune-related colitis, it deserves special attention. Camrelizumab is a programmed cell death 1 (PD-1) inhibitor that was developed by Jiangsu Hengrui Medicine Company. We reported the clinical data of a case of hepatocellular carcinoma with immune-related colitis after treatment with camrelizumab. A 63-year-old man with hepatocellular carcinoma developed diarrhea and hematochezia after receiving 4 cycles of camrelizumab. Endoscopy showed multiple flake congestion and edema in the terminal ileum and total colon mucosa with bright red surface. Pathological evaluation showed chronic inflammation of colonic mucosa. After giving 0.25g bid of enteric-coated sulfasalazine tablets orally for 6 weeks, his colitis improved. Camrelizumab can induce immune-related colitis. Sulfasalazine could be used to reduce adverse reactions of glucocorticoids.

The frequency and differential pleiotropy of phenotypic nonspecificity in Drosophila melanogaster.

The regulation of the initiation of transcription by transcription factors is often assumed to be dependent on specific recognition of DNA-binding sites and nonredundant. However, the redundant induction or rescue of a phenotype by transcription factors, phenotypic nonspecificity, challenges these assumptions. To assess the frequency of phenotypic nonspecificity in the rescue of transcription factor phenotypes, seven transcription factor phenotypes (labial, Deformed, Sex combs reduced, Ultrabithorax, fruitless, doublesex, and apterous) were screened for rescue by the expression of 12, or more, nonresident transcription factors. From 308 assessments of rescue by nonresident transcription factors, 18 rescues were identified across 6 of the 7 transcription factor phenotypes. Seventeen of the 18 rescues were with transcription factors that recognize distinct DNA-binding sites relative to the resident transcription factors. All rescues were nonuniform across pleiotropic transcription factor phenotypes suggesting extensive differential pleiotropy of the rescue. Primarily using RNAi to knockdown expression, and with the exceptions of the requirement of Bric a Brac 1 for female abdominal pigmentation and Myb oncogene-like for wing development, no evidence was found for a role of the other 16 nonresident transcription factor in the transcription factor phenotypes assessed. Therefore, these 16 rescues are likely due to functional complementation and not due to the expression of an epistatic function in the developmental/behavioral pathway. Phenotypic nonspecificity is both differentially pleiotropic and frequent, as on average 1 in 10-20 nonresident transcription factors rescue a phenotype. These observations will be important in future considerations of transcription factors function.

Single-port robot-assisted perineal radical prostatectomy with the da Vinci XI system: initial experience and learning curve using the cumulative sum method.

BACKGROUND: To evaluate the early functional and oncological outcomes of single-port robot-assisted perineal radical prostatectomy (sp-pRARP) using the da Vinci XI system and analyze its learning curve using the cumulative sum (CUSUM) method. METHODS: The clinical data of 50 patients who underwent sp-pRARP for localized prostate cancer between May 2020 and May 2022 in our center by a single surgeon were analyzed retrospectively. Demographic information, preoperative and postoperative variables, complications, early functional and oncological outcomes of patients were recorded. The CUSUM method was used to illustrate the learning curve based on operation time. RESULTS: All surgeries were completed without conversion. The median (interquartile range, IQR) operation time was 205.0 (82.5) min, whereas the median (IQR) docking time was 30.0 (15.0) min and the console time was 120.0 (80.5) min. The median (IQR) estimated blood loss (EBL) was 50.0 (137.5) mL. Positive surgical margins were detected in five patients (10.0%). The continence rate was 40.9%, 63.6%, 88.4%, and 97.7% at the 1, 3, 6, and 12 months after surgery. According to the CUSUM plot, the inflection points of the learning curve were 20 cases, splitting the case series into "early phase" and "late phase." In "late phase" cases, there was less time spent on each step of the operation and less EBL. CONCLUSIONS: Sp-pRARP using the da Vinci XI system was verified to be a feasible and reliable surgical approach. According to the CUSUM plot, 20 cases was considered the turning point for surgeons to master the novel technique.

[Consistency analysis of pepsin immunohistochemistry and pepsin test box in the diagnosis of laryngopharyngeal reflux].

Objective:To analyze the consistency of pepsin assay kit, pepsin IHC, reflux symptom index（RSI） and reflux finding score（RFS） in the diagnosis of laryngopharyngeal reflux disease（LPRD）. Methods:The clinical data of 61 inpatients with laryngeal diseases who were admitted to the Department of Otolaryngology, the First Affiliated Hospital of Kunming Medical University from May 2020 to December 2021 were retrospectively analyzed. The RSI and RFS scores, the Formwitz score of pepsin immunohistochemistry, and the results of pepsin detection kit were recorded. ICC group correlation coefficient and Kappa consistency analysis was used for three detection methods. Results:Among 61 patients, 30 cases were positive and 31 cases were negative for the pepsin test kit, with a positive rate of 49.18%. The positive rate of pepsin immunohistochemistry was 45.90%（28/61）, and the diagnostic agreement rate between the two was 70.49%. The consistency between them was high（κ=0.409）. The positive rate of RSI and RFS in diagnosing LPRD was 62.30%（38/61）, and the consistency rate was 73.77% with pepsin detection kit. The consistency between them was high（κ=0.486）. Taking pepsin IHC as the reference standard, the sensitivity, specificity, positive predictive value and negative predictive value of pepsin detection kit were 71.43%（20/28）, 69.70%（23/33）, 66.67%（20/30） and 74.19%（23/31）, respectively. Using RSI and RFS scales as reference criteria, the sensitivity, specificity, positive predictive value and negative predictive value of pepsin detection kit were 89.29%（25/28）, 60.61%（20/33）, 65.79%（25/38） and 86.96%（20/23）, respectively. Analysis of correlation coefficient within ICC group: ICC value was 0.628, 95% confidence interval（0.497-0.741）, the three methods have good consistency. Conclusion:The RSI and RFS scale scores were in good agreement with the pepsin test kit, and the pepsin test kit was also in good agreement with pepsin immunohistochemistry. As a non-invasive diagnostic technique, the pepsin test kit can be widely used in the diagnosis of pharyngeal reflux in combination with pepsin immunohistochemistry and RSI and RFS scale.