ObjectiveGastrodia elata has evolved ecological types with shortened rhizome internodes and diversified flower and fruit coloration in response to different altitudes. Studying the genetic mechanisms of different ecotype germplasm is significant for guiding variety breeding in different cultivation areas. MethodsThe bHLH gene family was identified based on the whole-genome datasets of G. elata f. elata and G. elata f. glauca. Subsequently， the gene family members were subject to analysis， including gene structure， chromosomal localization， cis-acting elements， gene synteny， and phylogeny. Combined with transcriptome data and quantitative Real-time PCR， the expression patterns of bHLH genes in the stems of the different G. elata ecotype germplasm were analyzed. Finally， correlation analysis was conducted between gene expression patterns and color to obtain the key bHLH genes regulating the color formation of stem. ResultsA total of 63 bHLH genes were identified in both G elata f. elata and G. elata f. glauca， unevenly distributed across 17 chromosomes and clustered into 16 subfamilies， with significant expansion in some family members. Obvious inversions of bHLH genes on the same chromosome and interchromosomal translocations were detected in the two ecotype germplasm. Among these genes， 12 bHLH genes （such as bHLH62-3 and bHLH74） were associated with the bright yellow color of G elata f. elata stem， while 9 bHLH genes （such as PIL13， UNE12， and bHLH130） were correlated with the red color of G. elata f. glauca stem. Compared to G. elata f. glauca， the bHLH48 expression level was significantly higher in flowers and scale leaves of G elata f. elata， and the bHLH62-3 expression level was significantly higher in all organs of G elata f. elata. ConclusionsFunctional pathway divergence of the bHLH family members has occurred across different chromosomes in G elata f. elata and G. elata f. glauca. Through synergism or antagonism with other genes， 21 bHLH genes participate in the coloration metabolic pathway regulation of stems， flowers， and fruits. Specifically， bHLH62-3 is involved in regulating stem color differentiation in the anthocyanin biosynthesis pathway of G. elata， thus relevant to the color formation of stem. Additionally， GebHLH48 positively regulates flowering-related pathways to promote the early-flowering phenotype of G. elata f. elata. These findings have laid the foundation for analyzing the genetic regulatory mechanisms underlying the color formation of the G. elata stem.

ObjectiveGastrodia elata has evolved ecological types with shortened rhizome internodes and diversified flower and fruit coloration in response to different altitudes. Studying the genetic mechanisms of different ecotype germplasm is significant for guiding variety breeding in different cultivation areas. MethodsThe bHLH gene family was identified based on the whole-genome datasets of G. elata f. elata and G. elata f. glauca. Subsequently， the gene family members were subject to analysis， including gene structure， chromosomal localization， cis-acting elements， gene synteny， and phylogeny. Combined with transcriptome data and quantitative Real-time PCR， the expression patterns of bHLH genes in the stems of the different G. elata ecotype germplasm were analyzed. Finally， correlation analysis was conducted between gene expression patterns and color to obtain the key bHLH genes regulating the color formation of stem. ResultsA total of 63 bHLH genes were identified in both G elata f. elata and G. elata f. glauca， unevenly distributed across 17 chromosomes and clustered into 16 subfamilies， with significant expansion in some family members. Obvious inversions of bHLH genes on the same chromosome and interchromosomal translocations were detected in the two ecotype germplasm. Among these genes， 12 bHLH genes （such as bHLH62-3 and bHLH74） were associated with the bright yellow color of G elata f. elata stem， while 9 bHLH genes （such as PIL13， UNE12， and bHLH130） were correlated with the red color of G. elata f. glauca stem. Compared to G. elata f. glauca， the bHLH48 expression level was significantly higher in flowers and scale leaves of G elata f. elata， and the bHLH62-3 expression level was significantly higher in all organs of G elata f. elata. ConclusionsFunctional pathway divergence of the bHLH family members has occurred across different chromosomes in G elata f. elata and G. elata f. glauca. Through synergism or antagonism with other genes， 21 bHLH genes participate in the coloration metabolic pathway regulation of stems， flowers， and fruits. Specifically， bHLH62-3 is involved in regulating stem color differentiation in the anthocyanin biosynthesis pathway of G. elata， thus relevant to the color formation of stem. Additionally， GebHLH48 positively regulates flowering-related pathways to promote the early-flowering phenotype of G. elata f. elata. These findings have laid the foundation for analyzing the genetic regulatory mechanisms underlying the color formation of the G. elata stem.

ObjectiveThe biosynthesis of heterophyllin B （HB）， a cyclopeptide from Pseudostellaria heterophylla， is regulated by various abiotic stresses. Elucidating the transcriptional regulatory mechanism underlying HB biosynthesis is of great guiding significance for the directional improvement of P. heterophylla varieties and the enhancement of HB content. MethodsBased on transcriptome data from different tissues of P. heterophylla， transcription factors （TFs） specifically upregulated and highly expressed in the phloem of tuberous roots were screened through a combination of Mfuzz time-series clustering， transcription factor family prediction， and correlation analysis. Quantitative real-time polymerase chain reaction （Real-time PCR） was employed to analyze expression patterns of candidate TFs under abscisic acid （ABA） induction， and the dual-luciferase reporter assay was applied to verify their regulatory effects on HB precursor genes. ResultsContent determination showed that HB accumulated at the highest in the phloem of P. heterophylla tuberous roots （34 μg·g^-1 fresh weight）. Transcriptome analysis identified 15 868 up-regulated differentially expressed genes （DEGs）， among which 4 375 were specifically up-regulated in the phloem. From these， 25 TFs highly expressed in the phloem were screened. Correlation analysis revealed that the expression level of WRKY70 was significantly positively correlated with HB content， the expression of precursor genes prePhHB， and PhPOP1. Additionally， WRKY70 expression was significantly upregulated under ABA induction. Dual-luciferase reporter assay confirmed that WRKY70 specifically activated the transcriptional activity of the prePhHB promoter （Pro-prePhHB）. ConclusionHB is mainly accumulated in the phloem of P. heterophylla tuberous roots. WRKY70 positively regulates HB biosynthesis by directly activating the promoter activity of prePhHB. This study clarifies the key role of WRKY70 in the regulation of HB biosynthesis and provides an important theoretical basis for the in-depth analysis of the molecular regulatory mechanism underlying HB biosynthesis.

BACKGROUND: Currently, lung cancer is one of the malignant tumors with a high morbidity and mortality all over the world. However, the exact mechanisms underlying lung cancer progression remain unclear. The tumor necrosis factor receptor associated factor (TRAF) family members are cytoplasmic adaptor proteins, which function as both adaptor proteins and ubiquitin ligases to regulate diverse receptor signalings, leading to the activation of nuclear factor kappa-B (NF-κB), mitogen-activated protein kinase (MAPK) and interferon regulatory factor (IRF) signaling. The aim of this study was to investigate the expression of TRAFs in different tissues and cancer types, as well as its mRNA expression, protein expression, prognostic significance and functional enrichment analysis in non-small cell lung cancer (NSCLC), in order to provide new strategies for the diagnosis and treatment of NSCLC. METHODS: RNA sequencing data from the The Genotype-Tissue Expression database was used to analyze the expression patterns of TRAF family members in different human tissues. RNA sequencing data from the Cancer Cell Line Encyclopedia database was used to analyze the expression patterns of TRAF family members in different types of cancer cell lines. RNA sequencing data from the The Cancer Genome Atlas (TCGA) database was used to analyze the mRNA levels of TRAF family members across different types of human cancers. Immunohistochemistry (IHC) analyses from HPA database were used to analyze the TRAF protein levels in NSCLC [lung adenocarcinoma (LUAD) and lung squamous carcinoma (LUSC)]. Overall survival analysis was performed by Log-rank test using original data from Kaplan-Meier Plotter database to evaluate the correlation between TRAF expressions and prognosis. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses were performed on the TRAF family-related genes using RNA sequencing data from the TCGA database for NSCLC. The correlation between the expression levels of TRAF family members and the tumor immune microenvironment was analyzed using the ESTIMATE algorithm based on RNA sequencing data from the TCGA database. RESULTS: The TRAF family members exhibited significant tissue-specific expression heterogeneity. TRAF2, TRAF3, TRAF6 and TRAF7 were widely expressed in most tissues, while the expressions of TRAF1, TRAF4 and TRAF5 were restricted to specific tissues. The expressions of TRAF family members were highly specific among different types of cancer cell lines. In mRNA database of LUAD and LUSC, the expressions of TRAF2, TRAF4, TRAF5 and TRAF7 were significantly upregulated; while TRAF6 did the opposite; moveover, TRAF1 and TRAF3 only displayed a significant upregulation in LUAD and LUSC, respectively. Except for TRAF3, TRAF4 and TRAF7, other TRAF proteins displayed an obviously deeper IHC staining in LUAD and LUSC tissues compared with normal tissues. Additionally, patients with higher expression levels of TRAF2, TRAF4 and TRAF7 had shorter overall survival; while patients with higher expression levels of TRAF3, TRAF5 and TRAF6 had significantly longer overall survival; however, no significant difference had been observed between TRAF1 expression and the overall survival. TRAF family members differentially regulated multiple pathways, including NF-κB, immune response, cell adhesion and RNA splicing. The expression levels of TRAF family members were closely associated with immune cell infiltration and stromal cell content in the tumor immune microenvironment, with varying positive and negative correlations among different members. CONCLUSIONS TRAF family members exhibit highly specific expression differences across different tissues and cancer types. Most TRAF proteins exhibit upregulation at both mRNA and protein levels in NSCLC, whereas, only upregulated expressions of TRAF2, TRAF4 and TRAF7 predict worse prognosis. The TRAF family members regulate processes such as inflammation, immunity, adhesion and splicing, and influence the tumor immune microenvironment.
Humans ; Carcinoma, Non-Small-Cell Lung/pathology* ; Lung Neoplasms/mortality* ; Prognosis ; Gene Expression Regulation, Neoplastic ; Tumor Necrosis Factor Receptor-Associated Peptides and Proteins/metabolism*

Background::T-cell-mediated immunity is crucial for the effective clearance of viral infection, but the T-cell-mediated immune responses that are induced by booster doses of inactivated severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccines in people living with human immunodeficiency virus (PLWH) remain unclear.Methods::Forty-five PLWH who had received antiretroviral therapy (ART) for more than two years and 29 healthy controls (HCs) at Beijing Youan Hospital were enrolled to assess the dynamic changes in T-cell responses between the day before the third vaccine dose (week 0) and 4 or 12 weeks (week 4 or week 12) after receiving the third dose of inactivated SARS-CoV-2 vaccine. Flow cytometry, enzyme-linked immunospot (ELISpot), and multiplex cytokines profiling were used to assess T-cell responses at the three timepoints in this study.Results::The results of the ELISpot and activation-induced marker (AIM) assays showed that SARS-CoV-2-specific T-cell responses were increased in both PLWH and HCs after the third dose of the inactivated SARS-CoV-2 vaccine, and a similar magnitude of immune response was induced against the Omicron (B.1.1.529) variant compared to the wild-type strain. In detail, spike-specific T-cell responses (measured by the ELISpot assay for interferon γ [IFN-γ] release) in both PLWH and HCs significantly increased in week 4, and the spike-specific T-cell responses in HCs were significantly stronger than those in PLWH 4 weeks after the third vaccination. In the AIM assay, spike-specific CD4 + T-cell responses peaked in both PLWH and HCs in week 12. Additionally, significantly higher spike-specific CD8 + T-cell responses were induced in PLWH than in HCs in week 12. In PLWH, the release of the cytokines interleukin-2 (IL-2), tumour necrosis factor-alpha (TNF-α), and IL-22 by peripheral blood mononuclear cells (PBMCs) that were stimulated with spike peptides increased in week 12. In addition, the levels of IL-4 and IL-5 were higher in PLWH than in HCs in week 12. Interestingly, the magnitude of SARS-CoV-2-specific T-cell responses in PLWH was negatively associated with the extent of CD8 + T-cell activation and exhaustion. In addition, positive correlations were observed between the magnitude of spike-specific T-cell responses (determined by measuring IFN-γ release by ELISpot) and the amounts of IL-4, IL-5, IL-2 and IL-17F. Conclusions::Our findings suggested that SARS-CoV-2-specific T-cell responses could be enhanced by the booster dose of inactivated COVID-19 vaccines and further illustrate the importance of additional vaccination for PLWH.

Objective:To analyze the clinical data of histiocytic necrotizing lymphadenitis(HNL),comparing the similarities and differences between children and adults,to deepen the understanding of the disease by clinical physicians,and to improve diagnostic rate and reduce misdiagnosis and mistreat-ment.Methods:The clinical data of hospitalized patients with histiocytic necrotizing lymphadenitis diagnosed by biopsy from January 2010 to August 2023 in Peking University First Hospital were collec-ted,and the clinical features,laboratory examinations,pathological features,treatments with antibiotics and glucocorticoids,and prognosis of histiocytic necrotic lymphadenitis were analyzed.Grouped based on age,the differences of clinical characteristics,laboratory tests,treatment,and prognosis between the children group(＜16 years old)and the adult group(≥16 years old)were compared.Results:Among the 81 enrolled patients,there were 42 males and 39 females.The median age was 21(14,29)years,the median duration of disease was 20.0(13.0,30.0)days,and the median length of hospital stay was 13.0(10.0,15.0)days.The first symptoms were fever,lymphadenopathy,and both.All the patients had enlarged lymph nodes with different parts and sizes,96.3％(78 of 81)of the patients had cervical lymphadenopathy,50.6％(41 of 81)had bilateral cervical lymphadenopathy,55.6％(45 of 81)had supraclavicular,axillary or inguinal lymphadenopathy,and the median lymph node diameter was 20.0(20.0,30.0)mm.Only one patient had no fever,the other 80 patients had fever,the median peak body temperature was 39.0(38.0,39.8)℃.Accompanying symptoms:rash(8.6％,7/81),fatigue(34.6％,28/81),night sweating(8.6％,7/81),chills(25.3％,25/81),muscle soreness(13.6％,11/81),and joint pain(6.2％,5/81).There were 17 cases(21.0％,17/81)of hepatosplenomegaly,of which 12 cases(70.6％,12/17)were splenomegaly.68.8％(55/80)of patients had a decrease in white blood cell(WBC)count,with 47.5％(38/80)increased in lymphocyte(LY)proportion,53.4％(39/73)increased in high-sensitivity C-reactive protein(CRP),79.2％(57/72)increased in erythrocyte sedimentation rate(ESR),22.2％(18/81)increased in alanine transaminase(ALT),27.2％(22/81)elevated in aspartate transaminase(AST),and 81.6％(62/76)elevated in lactate dehydrogenase(LDH).All the 81 patients underwent lymph node biopsy,and 77.8％(63/81)of the patients showed that most of the structures in the lymph nodes were destroyed or disappeared,and 16.0％(13/81)of the lymph nodes were still in existence,hyperplasia and normal lymph node were 1.2％(1/81)respectively,and 3.7％(3/81)had normal lymph node structures.Immunohistochemical staining was performed in 67 cases.The percentages of CD3+and CD68(KP1)+were respectively 97.0％(65/67),and MPO+were 94.0％(63/67).In the study,51 patients(63.0％,51/81)were treated with glucocorticoid therapy after diagnosis.The median time for temperature to return to normal was 1.0(1.0,4.0)days after glu-cocorticoid therapy.when the glucocorticoid treatment worked best,the body temperature could drop to normal on the same day.There were significant differences in length of stay,predisposing factors,chills,the rate of increase in high-sensitivity CRP,antibiotic and glucocorticoid treatment between the adults and children groups(P＜0.05).Conclusion:In clinical practice,if there are cases with unexplained fever,su-perficial lymph node enlargement,and reduced white blood cells as clinical characteristics,and general anti-biotics treatment is ineffective,histiocytic necrotic lymphadenitis should be considered.Lymph node biopsy should be performed as early as possible to clarify the diagnosis,reduce misdiagnosis and mistreatment,and symptomatic treatment should be the main treatment.Glucocorticoids therapy has a definite therapeutic effect.

Individuals with type 1 diabetes or advanced type 2 diabetes suffer insufficient insulin secretion, leading to symptoms of hyperglycemia. To maintain the normal blood glucose levels, those people with diabetes must administer insulin multiple times a day. However, insulin requirements are influenced by several factors such as diet, exercise, and illness, combined with the narrow therapeutic index, making accurate insulin dosage challenging. Excessive insulin administration can even pose life-threatening risks. In addition, frequent daily insulin injections place considerable physiological and psychological burdens on patients. To tackle these challenges, researchers have embarked on the development of closed-loop insulin delivery systems. These systems adjust insulin dosages in real-time changes based on the patient’s blood glucose levels, therefore enhancing both the safety and effectiveness of insulin therapy. This review categorizes closed-loop insulin delivery systems into two types: electronic-based and material-based systems, based on their compositional attributes. The exploration of both types covers their components, developmental history, clinical applications, current pros and cons, and future directions. The relative strengths and limitations of these two categories of closed-loop insulin delivery systems are also compared and discussed.

Methamphetamine (METH) abuse is associated with significant neurotoxicity, high addiction potential, and behavioral abnormalities. Recent studies have identified a connection between the gut microbiota and METH-induced neurotoxicity and behavioral disorders. However, the underlying causal mechanisms linking the gut microbiota to METH pathophysiology remain largely unexplored. In this study, we employed fecal microbiota transplantation (FMT) and antibiotic (Abx) intervention to manipulate the gut microbiota in mice administered METH. Furthermore, we supplemented METH-treated mice with short-chain fatty acids (SCFAs) and pioglitazone (Pio) to determine the protective effects on gut microbiota metabolism. Finally, we assessed the underlying mechanisms of the gut-brain neural circuit in vagotomized mice. Our data provide compelling evidence that modulation of the gut microbiome through FMT or microbiome knockdown by Abx plays a crucial role in METH-induced neurotoxicity, behavioral disorders, gut microbiota disturbances, and intestinal barrier impairment. Furthermore, our findings highlight a novel prevention strategy for mitigating the risks to both the nervous and intestinal systems caused by METH, which involves supplementation with SCFAs or Pio.

Background::Cluster of differentiation 8 (CD8 T) cells play critical roles in eradicating human immunodeficiency virus (HIV)-1 infection, but little is known about the effects of T cells expressing CD8 at low levels (CD8 low) or high levels (CD8 high) on HIV-1 replication inhibition after HIV-1 invasion into individual. Methods::Nineteen patients who had been acutely infected with HIV-1 (AHI) and 20 patients with chronic infection (CHI) for ≥2 years were enrolled in this study to investigate the dynamics of the quantity, activation, and immune responses of CD3 +CD8 low T cells and their counterpart CD3 +CD8 high T cells at different stages of HIV-1 infection. Results::Compared with healthy donors, CD3 +CD8 low T cells expanded in HIV-1-infected individuals at different stages of infection. As HIV-1 infection progressed, CD3 +CD8 low T cells gradually decreased. Simultaneously, CD3 +CD8 high T cells was significantly reduced in the first month of AHI and then increased gradually as HIV-1 infection progressed. The classical activation of CD3 +CD8 low T cells was highest in the first month of AHI and then reduced as HIV-1 infection progressed and entered the chronic stage. Meanwhile, activated CD38 -HLA-DR +CD8 low T cells did not increase in the first month of AHI, and the number of these cells was inversely associated with viral load ( r = -0.664, P = 0.004) but positively associated with the CD4 T-cell count ( r = 0.586, P = 0.014). Increased programmed cell death protein 1 (PD-1) abundance on CD3 +CD8 low T cells was observed from the 1st month of AHI but did not continue to be enhanced, while a significant T cell immunoreceptor with immunoglobulin and immunoreceptor tyrosine-based inhibition motif (ITIM) domains (TIGIT) abundance increase was observed in the 12th month of infection. Furthermore, increased PD-1 and TIGIT abundance on CD3 +CD8 low T cells was associated with a low CD4 T-cell count (PD-1: r = -0.456, P = 0.043; TIGIT: r = -0.488, P = 0.029) in CHI. Nonetheless, the nonincrease in PD-1 expression on classically activated CD3 +CD8 low T cells was inversely associated with HIV-1 viremia in the first month of AHI ( r = -0.578, P = 0.015). Notably, in the first month of AHI, few CD3 +CD8 low T cells, but comparable amounts of CD3 +CD8 high T cells, responded to Gag peptides. Then, weaker HIV-1-specific T-cell responses were induced in CD3 +CD8 low T cells than CD3 +CD8 high T cells at the 3rd and 12th months of AHI and in CHI. Conclusions::Our findings suggest that CD3 +CD8 low T cells play an anti-HIV role in the first month of infection due to their abundance but induce a weak HIV-1-specific immune response. Subsequently, CD3 +CD8 low T-cell number decreased gradually as infection persisted, and their anti-HIV functions were inferior to those of CD3 +CD8 high T cells.

Objective:To investigate the risk factors of surgical site infection (SSI) after colorectal surgery, and to establish and validate a risk prediction model nomogram.Methods:An observational study was conducted to retrospectively collect data of 6527 patients aged ≥16 years who underwent colorectal surgery in 56 domestic hospitals from March 1, 2021 to February 28, 2022 from the national Surgical Site Infection Surveillance network. The incidence of SSI after surgery was 2.3% (149/6527). According to the ratio of 7:3, 6527 patients were randomly divided into the modeling cohort (4568 cases) and the validation cohort (1959 cases), and there was no statistically significant difference between the two datasets ( P>0.05). Univariate analysis was performed using t test /Mann-Whitney U test /χ 2 test. Multivariate analysis was performed using binary logistic regression to establish a preliminary model and select variables using Lasso analysis to establish an optimized model nomogram. The discrimination and calibration of the model were evaluated by ROC curve, calibration curve, and Hosmer-Lemeshow test. AUC value>0.7 is considered a good discrimination of the model. The Bootstrap method (repeated self-sampling 1000 times) was used to verify the constructed model internally and externally to evaluate the accuracy of the constructed model. Results:Multivariate analysis showed that history of chronic liver disease (OR=3.626, 95%CI: 1.297-10.137, P<0.001) and kidney disease (OR=1.567,95%CI:1.042-2.357, P=0.038), surgical antibiotic prophylaxis (OR=1.564, 95%CI:1.038-2.357, P=0.035), and emergency surgery (OR=1.432,95%CI: 1.089-1.885, P=0.021), open surgery (OR=1.418, 95%CI:1.045-1.924, P=0.042), preoperative stoma (OR=3.310, 95%CI:1.542-7.105, P<0.001), postoperative stoma (OR=2.323,95%CI: 1.537-8.134, P<0.001), surgical incision type above grade II (OR=1.619,95%CI:1.097-2.375, P=0.014), and each unit increase in total bilirubin (OR=1.003,95%CI:-0.994-1.012, P=0.238), alanine aminotransferase (OR=1.006, 95%CI:1.001-1.011, P=0.032), blood urea nitrogen (OR=1.003,95%CI:0.995-1.011, P=0.310), blood glucose (OR=1.024, 95%CI:1.005-1.043, P=0.027), C-reactive protein (OR=1.007, 95%CI:1.003-1.011, P<0.001), length of incision (OR=1.042, 95%CI:1.002-1.087, P=0.031), surgical duration (OR=1.003,95%CI:1.001-1.005, P=0.017), and surgical blood loss (OR=1.001,95%CI: 1.000-1.002, P=0.045) were risk factors for SSI after colorectal surgery. Each unit increase in albumin level (OR=0.969,95%CI:0.941-0.998, P=0.036) was an independent protective factor for SSI after colorectal surgery. The area under the curve of the optimized model obtained by internal and external validation were 0.768 (95%CI: 0.723-0.813) and 0.753 (95%CI: 0.680-0.832), respectively. The predicted value of the calibration curve was basically consistent with the actual value. Conclusions:The risk prediction model for SSI after colorectal surgery constructed in this study has good discrimination and calibration. The nomogram created in this model can provide an evaluation basis for the observed rate and expected event rate of SSI after clinical colorectal surgery.