Decompression sickness-induced skeletal muscle injury: an animal model and pathological analysis.

Aims: The primary objective of this investigation is to establish an animal model that accurately represents skeletal muscle injury as a consequence of decompression sickness. Additionally, this study aims to delineate the potential mechanisms underlying the development and progression of skeletal muscle damage associated with decompression sickness. Materials and methods: (1) In this research, rats were utilized as experimental models and subjected to 600 kPa pressure in an air medium for a duration of 60 min, followed by decompression at a consistent rate of 1.5 min to reach atmospheric pressure in order to establish an animal model for decompression injury. Assessment of decompression injury involved the observation of general symptoms and signs, as well as histopathological examination of lung tissue to determine the extent of damage in the pulmonary system of rats. (2) Building on the rat decompression injury model, we conducted pathological and serological examinations to assess the status of rat skeletal muscle. Additionally, we investigated the signaling mechanism of the TLR9-MyD88 pathway in mediating alterations in rat skeletal muscle resulting from decompression injury, and evaluated the effects of decompression injury on apoptosis in rat skeletal muscle. Results: Repeated decompression induces significant damage to skeletal muscle tissue, characterized by edema, fiber rupture, and atrophy. This process also leads to a transient elevation in creatine kinase (CK-MM) levels in rat serum, as well as an upregulation of proteins such as TLR9, MyD88, p38, and ERK in rat skeletal muscle tissue. Furthermore, repeated decompression results in a temporary increase in the transcription levels of Atrogen-1mRNA and MuRF-1mRNA in rat skeletal muscle tissue. Discussion: The decompression protocol applied in this study successfully induced decompression sickness in a rat model, leading to skeletal muscle damage that was consistent with the expected pathology of decompression injury. Despite the initial injury, the rats showed evidence of adaptation following prolonged exposure to decompression conditions.

Lanthanide MOF-based luminescent sensor array for detection and identification of contaminants in water and biomarkers.

In today's society, heavy metal ions and antibiotic contaminants have caused great harm to water systems and human health. In this study, six isostructural lanthanide metal-organic frameworks [Ln(H3imda)2(TPA)(H2O)2](Tb for CUST-881, Eu for CUST-882, Dy for CUST-883, Er for CUST-884, Nd for CUST-885, Sm for CUST-886) were constructed by selecting terephthalic acid (TPA) and 4,5-Imidazoledicarboxylic acid (H3imda) and lanthanide metal ions via solvethermal method. Among them, CUST-881 and CUST-882 can selectively detect Fe3+, Cr2O72-, CrO42, and ceftriaxone sodium (CRO) in water systems and uric acid in urine. CUST-881 shows very low detection limits for these five substances. Furthermore, Principal Component Analysis (PCA) was used to distinguish Fe3+, Cr2O72-, CrO42-, and CRO in water. To our knowledge, this is the first time that they have been able to be simultaneously distinguished. In addition, the possible sensing mechanism was studied through UV-visible spectroscopy, Infrared spectroscopy, and PXRD analysis. Furthermore, the probe also showed satisfactory repeatability and recovery when applied to UA samples that simulated urine. Based on the above results, lanthanide metal-organic frameworks have great potential for practical monitoring of contaminants in water environments.

New findings on the incidence and management of CNS adverse reactions in ALK-positive NSCLC with lorlatinib treatment.

To explore the presentation and control of CNS adverse reactions in patients with ALK-positive NSCLC treated with lorlatinib. This study includes a retrospective case report from Sir Run Run Shaw Hospital on a lorlatinib-treated patient with CNS adverse reactions and a systematic literature review of similar cases until January 2023. The report detailed a case of a 74-year-old male with Grade III CNS adverse reactions 25 days after starting lorlatinib, which were reversible with dose modification and pharmacotherapy. The review indicated a 19.39% occurrence rate of such reactions, with a 17% improvement rate post-dose adjustment. CNS adverse reactions frequently occur in ALK-positive NSCLC patients on lorlatinib, yet they are reversible with appropriate management. Research should continue to optimize treatment protocols to decrease these reactions' frequency.

Extended Criteria Donor Use in Lung Transplants From Donation After Controlled Circulatory Death.

BACKGROUND: Most institutions apply the criteria for controlled donation after cardiac death (cDCD) lung retrieval identical to the criteria for donation after brain death (DBD). The availability of extended criteria donor (ECD) in lung transplants from cDCD remains unclear. METHODS: The United Network for Organ Sharing (UNOS) database was queried for adult lung transplants from cDCD, from May 03, 2005, to March 15, 2022. ECDs were defined by one or more items at variance from standard criteria: age 55 years or more, PaO2:FiO2 300 or less, smoking 20 pack-years or more, diabetes, or purulent secretions upon bronchoscopy. Recipients were divided into the standard criteria donor (SCD) group and the ECD group, and assessed for short- and long-term survival and postoperative events. RESULTS: Among 827 records, the SCD and ECD group showed no differences in 5-year (P = .56) survival. No significant differences were found in 30-day, 90-day, 1-year mortality and postoperative outcomes before discharge, whether in length of hospital stay, rate of ventilator support for >48 hours or reintubation, incidence of grade 3 PGD 72 hours posttransplant, acute rejection, or dialysis. None of the 5 donor factors used as criteria for lung retrieval was independently associated with cDCD recipient survival. CONCLUSIONS: Using donor lungs that extend the DBD criteria may be a safe strategy in cDCD lung transplantation. However, the current criteria may not be a perfect fit for cDCD lung retrieval. The specific DCD criteria for standard lung retrieval need to be determined.

Fluorescent Sensors Based on Lanthanide-Based Metal-Organic Frameworks via Devices and pH Response.

In light of the escalating industrial and environmental pollution, there is a pressing need for the development of novel materials capable of swiftly detecting pollutants. Here, we report the synthesis of five lanthanide metal-organic frameworks sharing a common structure, prepared via a hydrothermal method and denoted as [Ln2(H2DHBDC)2(phen)(H2O)6]n (where CUST-888 corresponds to Tb, CUST-889 corresponds to Eu, CUST-890 corresponds to Gd, CUST-891 corresponds to Dy, and CUST-892 corresponds to Nd). Notably, CUST-888 and CUST-889 exhibit discernible visual alterations in response to acidic and alkaline conditions. To assess their practical utility, luminescent test strips and light-emitting diode lights based on CUST-888 and CUST-889 were devised, enabling the visual detection of luminescence color changes induced by Hg2+, Cr2O72-, tetracycline, and 2,4,6-trinitrophenol. Furthermore, highlighters derived from CUST-888 and CUST-889 were designed, showcasing robust stability, adjustable color, and substantial potential for application in the realm of anticounterfeiting.

Gene Therapy for Retinitis Pigmentosa: Current Challenges and New Progress.

Retinitis pigmentosa (RP) poses a significant threat to eye health worldwide, with prevalence rates of 1 in 5000 worldwide. This genetically diverse retinopathy is characterized by the loss of photoreceptor cells and atrophy of the retinal pigment epithelium. Despite the involvement of more than 3000 mutations across approximately 90 genes in its onset, finding an effective treatment has been challenging for a considerable time. However, advancements in scientific research, especially in gene therapy, are significantly expanding treatment options for this most prevalent inherited eye disease, with the discovery of new compounds, gene-editing techniques, and gene loci offering hope for more effective treatments. Gene therapy, a promising technology, utilizes viral or non-viral vectors to correct genetic defects by either replacing or silencing disease-causing genes, potentially leading to complete recovery. In this review, we primarily focus on the latest applications of gene editing research in RP. We delve into the most prevalent genes associated with RP and discuss advancements in genome-editing strategies currently employed to correct various disease-causing mutations.

Remodeling Intestinal Microbiota Alleviates Severe Combined Hyperlipidemia-Induced Nonalcoholic Steatohepatitis and Atherosclerosis in LDLR-/- Hamsters.

Combined hyperlipidemia (CHL) manifests as elevated cholesterol and triglycerides, associated with fatty liver and cardiovascular diseases. Emerging evidence underscores the crucial role of the intestinal microbiota in metabolic disorders. However, the potential therapeutic viability of remodeling the intestinal microbiota in CHL remains uncertain. In this study, CHL was induced in low-density lipoprotein receptor-deficient (LDLR-/-) hamsters through an 8-week high-fat and high-cholesterol (HFHC) diet or a 4-month high-cholesterol (HC) diet. Placebo or antibiotics were administered through separate or cohousing approaches. Analysis through 16S rDNA sequencing revealed that intermittent antibiotic treatment and the cohousing approach effectively modulated the gut microbiota community without impacting its overall abundance in LDLR-/- hamsters exhibiting severe CHL. Antibiotic treatment mitigated HFHC diet-induced obesity, hyperglycemia, and hyperlipidemia, enhancing thermogenesis and alleviating nonalcoholic steatohepatitis (NASH), concurrently reducing atherosclerotic lesions in LDLR-/- hamsters. Metabolomic analysis revealed a favorable liver lipid metabolism profile. Increased levels of microbiota-derived metabolites, notably butyrate and glycylglycine, also ameliorated NASH and atherosclerosis in HFHC diet-fed LDLR-/- hamsters. Notably, antibiotics, butyrate, and glycylglycine treatment exhibited protective effects in LDLR-/- hamsters on an HC diet, aligning with outcomes observed in the HFHC diet scenario. Our findings highlight the efficacy of remodeling gut microbiota through antibiotic treatment and cohousing in improving obesity, NASH, and atherosclerosis associated with refractory CHL. Increased levels of beneficial microbiota-derived metabolites suggest a potential avenue for microbiome-mediated therapies in addressing CHL-associated diseases.

FOXO1 is a master regulator of memory programming in CAR T cells.

A major limitation of chimeric antigen receptor (CAR) T cell therapies is the poor persistence of these cells in vivo1. The expression of memory-associated genes in CAR T cells is linked to their long-term persistence in patients and clinical efficacy2-6, suggesting that memory programs may underpin durable CAR T cell function. Here we show that the transcription factor FOXO1 is responsible for promoting memory and restraining exhaustion in human CAR T cells. Pharmacological inhibition or gene editing of endogenous FOXO1 diminished the expression of memory-associated genes, promoted an exhaustion-like phenotype and impaired the antitumour activity of CAR T cells. Overexpression of FOXO1 induced a gene-expression program consistent with T cell memory and increased chromatin accessibility at FOXO1-binding motifs. CAR T cells that overexpressed FOXO1 retained their function, memory potential and metabolic fitness in settings of chronic stimulation, and exhibited enhanced persistence and tumour control in vivo. By contrast, overexpression of TCF1 (encoded by TCF7) did not enforce canonical memory programs or enhance the potency of CAR T cells. Notably, FOXO1 activity correlated with positive clinical outcomes of patients treated with CAR T cells or tumour-infiltrating lymphocytes, underscoring the clinical relevance of FOXO1 in cancer immunotherapy. Our results show that overexpressing FOXO1 can increase the antitumour activity of human CAR T cells, and highlight memory reprogramming as a broadly applicable approach for optimizing therapeutic T cell states.

Depletion of ApoA5 aggravates spontaneous and diet-induced nonalcoholic fatty liver disease by reducing hepatic NR1D1 in hamsters.

Background: ApoA5 mainly synthesized and secreted by liver is a key modulator of lipoprotein lipase (LPL) activity and triglyceride-rich lipoproteins (TRLs). Although the role of ApoA5 in extrahepatic triglyceride (TG) metabolism in circulation has been well documented, the relationship between ApoA5 and nonalcoholic fatty liver disease (NAFLD) remains incompletely understood and the underlying molecular mechanism still needs to be elucidated. Methods: We used CRISPR/Cas9 gene editing to delete Apoa5 gene from Syrian golden hamster, a small rodent model replicating human metabolic features. Then, the ApoA5-deficient (ApoA5-/-) hamsters were used to investigate NAFLD with or without challenging a high fat diet (HFD). Results: ApoA5-/- hamsters exhibited hypertriglyceridemia (HTG) with markedly elevated TG levels at 2300 mg/dL and hepatic steatosis on a regular chow diet, accompanied with an increase in the expression levels of genes regulating lipolysis and small adipocytes in the adipose tissue. An HFD challenge predisposed ApoA5-/- hamsters to severe HTG (sHTG) and nonalcoholic steatohepatitis (NASH). Mechanistic studies in vitro and in vivo revealed that targeting ApoA5 disrupted NR1D1 mRNA stability in the HepG2 cells and the liver to reduce both mRNA and protein levels of NR1D1, respectively. Overexpression of human NR1D1 by adeno-associated virus 8 (AAV8) in the livers of ApoA5-/- hamsters significantly ameliorated fatty liver without affecting plasma lipid levels. Moreover, restoration of hepatic ApoA5 or activation of UCP1 in brown adipose tissue (BAT) by cold exposure or CL316243 administration could significantly correct sHTG and hepatic steatosis in ApoA5-/- hamsters. Conclusions: Our data demonstrate that HTG caused by ApoA5 deficiency in hamsters is sufficient to elicit hepatic steatosis and HFD aggravates NAFLD by reducing hepatic NR1D1 mRNA and protein levels, which provides a mechanistic link between ApoA5 and NAFLD and suggests the new insights into the potential therapeutic approaches for the treatment of HTG and the related disorders due to ApoA5 deficiency in the clinical trials in future.

Regulation of immune signal integration and memory by inflammation-induced chromosome conformation.

3-dimensional (3D) genome conformation is central to gene expression regulation, yet our understanding of its contribution to rapid transcriptional responses, signal integration, and memory in immune cells is limited. Here, we study the molecular regulation of the inflammatory response in primary macrophages using integrated transcriptomic, epigenomic, and chromosome conformation data, including base pair-resolution Micro-Capture C. We demonstrate that interleukin-4 (IL-4) primes the inflammatory response in macrophages by stably rewiring 3D genome conformation, juxtaposing endotoxin-, interferon-gamma-, and dexamethasone-responsive enhancers in close proximity to their cognate gene promoters. CRISPR-based perturbations of enhancer-promoter contacts or CCCTC-binding factor (CTCF) boundary elements demonstrated that IL-4-driven conformation changes are indispensable for enhanced and synergistic endotoxin-induced transcriptional responses, as well as transcriptional memory following stimulus removal. Moreover, transcriptional memory mediated by changes in chromosome conformation often occurred in the absence of changes in chromatin accessibility or histone modifications. Collectively, these findings demonstrate that rapid and memory transcriptional responses to immunological stimuli are encoded in the 3D genome.

Electroacupuncture attenuates nociceptive behaviors in a mouse model of cancer pain.

Pain is a major symptom in cancer patients, and cancer-induced bone pain (CIBP) is the most common type of moderate and severe cancer-related pain. The current available analgesic treatments for CIBP have adverse effects as well as limited therapeutic effects. Acupuncture is proved effective in pain management as a safe alternative therapy. We evaluated the analgesic effect of acupuncture in treatment of cancer pain and try to explore the underlying analgesic mechanisms. Nude mice were inoculated with cancer cells into the left distal femur to establish cancer pain model. Electroacupuncture (EA) treatment was applied for the xenograft animals. Pain behaviors of mice were evaluated, followed by the detections of neuropeptide-related and inflammation-related indicators in peripheral and central levels. EA treatment alleviated cancer-induced pain behaviors covering mechanical allodynia, thermal hyperalgesia and spontaneous pain, and also down-regulated immunofluorescence expressions of neuropeptide CGRP and p75 in the skin of affected plantar area in xenograft mice, and inhibited expressions of overexpressed neuropeptide-related and inflammation-related protein in the lumbar spinal cord of xenograft mice. Overall, our findings suggest that EA treatment ameliorated cancer-induced pain behaviors in the mouse xenograft model of cancer pain, possibly through inhibiting the expressions of neuropeptide-related and inflammation-related protein in central level following tumor cell xenografts.

Comparing the efficacy and safety of three surgical approaches for total hysterectomy (TSATH): protocol for a multicentre, single-blind, parallel-group, randomised controlled trial.

INTRODUCTION: Hysterectomy is the most common surgical procedure in the field of gynaecology. The traditional multiport laparoscopy, transumbilical laparoendoscopic single-site surgery (TU-LESS) and transvaginal natural orifice transluminal endoscopic surgery (vNOTES) hysterectomy approaches have been implemented to varying degrees in clinical practice. At present, although their feasibility has been proven, there are no large randomised controlled studies on postoperative rehabilitation. This study aims to evaluate postoperative recovery and assess the safety and effectiveness of these three surgical approaches for total laparoscopic hysterectomy. METHOD AND ANALYSIS: This is a multicentre, randomised, single-blind, three-arm, parallel-group, interventional clinical trial. Recruitment will be carried out in five tertiary hospitals in China. Patients diagnosed with benign uterine disease or precancerous lesions will be assigned to the vNOTES group, TU-LESS group and conventional laparoscopy group at a 1:1:1 ratio. The achievement rate of comprehensive indices of enhanced recovery after surgery (ERAS) within 24 hours postoperatively will be considered the primary outcome (the comprehensive indicators of ERAS include fluid intake, passing flatus, urination after catheter removal, ambulation and a Visual Analogue Scale score ≤3.) This study will use a non-inferiority test, with a power (1-ß) of 80% and a margin of -0.15, at a one-sided α of 0.0125. The sample size will be 480 patients (including an assumed 15% dropout rate), calculated according to the primary outcome. ETHICS AND DISSEMINATION: This study was approved on 25 April 2022 by the Medical Ethics Committee of West China Second University Hospital (2022(057)), Sichuan University, Chengdu, China. All participants will be required to provide informed consent before their participation in the study. The results of the trial will be submitted for publication in a peer-reviewed journal and presented at international conferences. PROTOCOL VERSION: V.3.0, 31 August 2023. TRIAL REGISTRATION NUMBER: ChiCTR2200057405.

ZIF-derived sulfides with tremella-like core-shell structure for high performance supercapacitors.

Metal-organic frameworks (MOFs) have emerged as promising active electrode materials in supercapacitors for its controllable porous structure and excellent physio-chemical properties. However, the poor conductivities keep it from achieving its full capacitance potential, which greatly limits its practical application. Here, a facile pathway is reported to fabricate the GO/Ni2ZnS4@NiCo2S4 composite with large specific surface area and favorable electrical conductivity. Thanks to the novel tremella-like core-shell structure and high-efficient synergistic effects among multi-components, the designed GO/Ni2ZnS4@NiCo2S4 electrode shows a high specific capacitance of 2284 F/g at 1 A/g. Furthermore, the asymmetric supercapacitor fabricated by coupling GO/Ni2ZnS4@NiCo2S4 positive electrode with biological carbon negative electrode achieves a remarkable energy density of 120 Wh kg-1 at a power density of 750 W kg-1.

High-fibre diets regulate antioxidative capacity and promote intestinal health by regulating bacterial microbiota in growing pigs.

This experiment was conducted to investigate the effects of a high-fibre diet on growth performance, nutrients digestibility, intestinal health, and intestinal microbiota composition of growing pigs. Twelve healthy "Duroc × Landrace × Yorkshire" castrates (49 ± 1.35 kg) were randomly divided into two groups with six replicates and one pig per replicate. The two diet treatments were fed the basal diet (CON) based on corn and soybean meal and high fibre diet (HF) respectively. The nutritional levels of the two treatments were the same. The experiment lasted 28 days. The results showed that the addition of 16% wheat bran fibre to the diet of growing pigs did not affect growth performance (p > 0.05). Compared with the CON, contents of isobutyric and butyric acid, GSH-PX and T-AOC in serum were increased in the HF. It decreased the gross energy digestibility and acetic acid content in feces of growing pigs (p < 0.05), the contents of GSH-PX and T-AOC in serum. It decreased the gross energy digestibility and acetic acid content in feces of growing pigs (p < 0.05). Compared with the CON, the Shannon, and Chao1 indexes of the HF were increased (p < 0.05). At the phylum level, the abundance of g_Lactobacillus increased in the HF (p < 0.05). Correlation analysis showed that a total of 18 microbial genera were correlated with antioxidant capacity and volatile fatty acid levels (p < 0.05). In summary, this study showed that adding 16% wheat bran to the diet of growing pigs had no effect on growth performance but helped to improve the richness and stability of intestinal microbiota, promote posterior intestinal fermentation and increase serum antioxidant capacity.

FOXO1 is a master regulator of CAR T memory programming.

Poor CAR T persistence limits CAR T cell therapies for B cell malignancies and solid tumors1,2. The expression of memory-associated genes such as TCF7 (protein name TCF1) is linked to response and long-term persistence in patients3-7, thereby implicating memory programs in therapeutic efficacy. Here, we demonstrate that the pioneer transcription factor, FOXO1, is responsible for promoting memory programs and restraining exhaustion in human CAR T cells. Pharmacologic inhibition or gene editing of endogenous FOXO1 in human CAR T cells diminished the expression of memory-associated genes, promoted an exhaustion-like phenotype, and impaired antitumor activity in vitro and in vivo. FOXO1 overexpression induced a gene expression program consistent with T cell memory and increased chromatin accessibility at FOXO1 binding motifs. FOXO1-overexpressing cells retained function, memory potential, and metabolic fitness during settings of chronic stimulation and exhibited enhanced persistence and antitumor activity in vivo. In contrast, TCF1 overexpression failed to enforce canonical memory programs or enhance CAR T cell potency. Importantly, endogenous FOXO1 activity correlated with CAR T and TIL responses in patients, underscoring its clinical relevance in cancer immunotherapy. Our results demonstrate that memory reprogramming through FOXO1 can enhance the persistence and potency of human CAR T cells and highlights the utility of pioneer factors, which bind condensed chromatin and induce local epigenetic remodeling, for optimizing therapeutic T cell states.

[Effect of electroacupuncture on myocardial fibrosis in spontaneously hypertensive rats based on cholinergic anti-inflammatory pathway].

OBJECTIVE: To observe the effect of electroacupuncture (EA) at "Neiguan" (PC 6) on myocardial fibrosis in spontaneously hypertensive rats (SHR), and explore preliminarily the mediating role of cholinergic anti-inflammatory pathway (CAP) and its downstream nuclear factor κB (NF-κB) signaling pathway. METHODS: Six 12-week-old WKY male rats were employed as the normal group. Eighteen 12-week-old SHR were randomly divided into 3 groups, i.e. a model group, an EA group and a blocking group (EA after blocking α7 nicotinic acetylcholine receptor [α7nAchR]), with 6 rats in each one. In the EA group, EA was delivered at "Neiguan"(PC 6) and the site 0.5 cm from its left side, with disperse-dense wave, 2 Hz/15 Hz in frequency and 1 mA in current intensity. One intervention took 30 min and was given once every 2 days, lasting 8 weeks. In the blocking group, prior to each EA, the α7nAchR specific blocker, α-bungartoxin was injected intravenously in the tails of the rats. After EA intervention, the systolic blood pressure (SBP), the diastolic blood pressure (DBP) and the mean arterial pressure (MAP) were measured with non-invasive blood pressure monitor. Using echocardiogram, the left ventricular (LV) anterior wall end-diastolic thickness (LVAWd) , LV posterior wall end-diastolic thickness (LVPWd) and the LV end-diastolic internal diameter (LVIDd) were measured. The level of hydroxyproline (Hyp) in the myocardial tissue was determined by using alkaline hydrolysis, and that of acetylcholine (Ach) was detected by ELISA. With the real-time PCR adopted, the mRNA expression of NF-κB p65, tumor necrosis factor α (TNF-α), interleukin (IL)-1ß and IL-6 were determined. RESULTS: Compared with the normal group, SBP, DBP, MAP, LVAWd and LVPWd were increased (P<0.01), and LVIDd was decreased (P<0.01) in the rats of the model group. SBP, DBP, MAP and LVAWd were dropped (P<0.01, P<0.05), and LVIDd rose (P<0.01) in the EA group when compared with those in the model group. The differences in the above indexes were not statistically significant between the blocking group and the model group (P>0.05). Compared with the normal group, Hyp level and the mRNA expression of NF-κB p65, TNF-α, IL-1ß and IL-6 in the myocardial tissue increased (P<0.01, P<0.05) and Ach level decreased (P<0.01) in the model group. Hyp level, the mRNA expression of NF-κB p65, TNF-α, IL-1ß and IL-6 in the myocardial tissue were reduced (P<0.05, P<0.01) and Ach level rose (P<0.01) in the EA group when compared with those in the model group. These indexes were not different statistically between the blocking group and the model group (P>0.05). CONCLUSION: CAP may be involved in ameliorating the pathological damage of myocardial fibrosis during EA at "Neiguan"(PC 6). The underlying effect mechanism is associated with up-regulating the neurotransmitter, Ach and down-regulating mRNA expression of NF-κB p65 and pro-inflammatory factors such as TNF-α, IL-1ß and IL-6 in myocardial tissue.

Modular pooled discovery of synthetic knockin sequences to program durable cell therapies.

Chronic stimulation can cause T cell dysfunction and limit the efficacy of cellular immunotherapies. Improved methods are required to compare large numbers of synthetic knockin (KI) sequences to reprogram cell functions. Here, we developed modular pooled KI screening (ModPoKI), an adaptable platform for modular construction of DNA KI libraries using barcoded multicistronic adaptors. We built two ModPoKI libraries of 100 transcription factors (TFs) and 129 natural and synthetic surface receptors (SRs). Over 30 ModPoKI screens across human TCR- and CAR-T cells in diverse conditions identified a transcription factor AP4 (TFAP4) construct that enhanced fitness of chronically stimulated CAR-T cells and anti-cancer function in vitro and in vivo. ModPoKI's modularity allowed us to generate an ∼10,000-member library of TF combinations. Non-viral KI of a combined BATF-TFAP4 polycistronic construct enhanced fitness. Overexpressed BATF and TFAP4 co-occupy and regulate key gene targets to reprogram T cell function. ModPoKI facilitates the discovery of complex gene constructs to program cellular functions.

[Development and Validation of A Prognostic Nomogram to Guide Decision-making 
in Lung Large Cell Neuroendocrine Carcinoma].

BACKGROUND: Lung large cell neuroendocrine carcinoma (LCNEC) is a rare and highly malignant lung tumor with a poor prognosis. Currently, most research on LCNEC is based on retrospective studies and lacks validation in the real world. The study aims to identify independent risk factors and establish and validate a predictive model for the prognosis of LCNEC. METHODS: Patient data were extracted from Surveillance, Epidemiology, and End Results (SEER) and our department's hospitalization records from 2010 to 2015 and 2016 to 2020, respectively. Kaplan-Meier analysis was used to evaluate overall survival (OS). OS is defined as the time from diagnosis to death or last follow-up for a patient. Univariate and multivariate Cox regression analyses were performed to identify significant prognostic factors and construct a Nomogram for predicting the prognosis of LCNEC. RESULTS: In total, 1892 LCNEC patients were included and divided into a training cohort (n=1288) and two validation cohorts (n=552, n=52). Univariate Cox regression analysis showed that age, gender, primary tumor site, laterality, T stage, N stage, M stage, surgery, and radiotherapy were factors that could affect the prognosis of LCNEC patients (P<0.05). Multivariate Cox analysis indicated that age, gender, primary tumor site, T stage, N stage, M stage, surgery, and radiotherapy were independent risk factors for the prognosis of LCNEC patients (P<0.05). Calibration curves and the concordance index (internal: 0.744±0.015; external: 0.763±0.020, 0.832±0.055) demonstrated good predictive performance of the model. CONCLUSIONS: Patients aged ≥65 years, male, with advanced tumor-node-metastasis (TNM) staging, and who have not undergone surgery or radiotherapy have a poor prognosis. Nomogram can provide a certain reference for personalized clinical decision-making for patients.