Identifying the risk factors of ICU-acquired fungal infections: clinical evidence from using machine learning.

Background: Fungal infections are associated with high morbidity and mortality in the intensive care unit (ICU), but their diagnosis is difficult. In this study, machine learning was applied to design and define the predictive model of ICU-acquired fungi (ICU-AF) in the early stage of fungal infections using Random Forest. Objectives: This study aimed to provide evidence for the early warning and management of fungal infections. Methods: We analyzed the data of patients with culture-positive fungi during their admission to seven ICUs of the First Affiliated Hospital of Chongqing Medical University from January 1, 2015, to December 31, 2019. Patients whose first culture was positive for fungi longer than 48 h after ICU admission were included in the ICU-AF cohort. A predictive model of ICU-AF was obtained using the Least Absolute Shrinkage and Selection Operator and machine learning, and the relationship between the features within the model and the disease severity and mortality of patients was analyzed. Finally, the relationships between the ICU-AF model, antifungal therapy and empirical antifungal therapy were analyzed. Results: A total of 1,434 cases were included finally. We used lasso dimensionality reduction for all features and selected six features with importance ≥0.05 in the optimal model, namely, times of arterial catheter, enteral nutrition, corticosteroids, broadspectrum antibiotics, urinary catheter, and invasive mechanical ventilation. The area under the curve of the model for predicting ICU-AF was 0.981 in the test set, with a sensitivity of 0.960 and specificity of 0.990. The times of arterial catheter (p = 0.011, OR = 1.057, 95% CI = 1.053-1.104) and invasive mechanical ventilation (p = 0.007, OR = 1.056, 95%CI = 1.015-1.098) were independent risk factors for antifungal therapy in ICU-AF. The times of arterial catheter (p = 0.004, OR = 1.098, 95%CI = 0.855-0.970) were an independent risk factor for empirical antifungal therapy. Conclusion: The most important risk factors for ICU-AF are the six time-related features of clinical parameters (arterial catheter, enteral nutrition, corticosteroids, broadspectrum antibiotics, urinary catheter, and invasive mechanical ventilation), which provide early warning for the occurrence of fungal infection. Furthermore, this model can help ICU physicians to assess whether empiric antifungal therapy should be administered to ICU patients who are susceptible to fungal infections.

Application status and optimization suggestions of tumor organoids and CAR-T cell co-culture models.

Tumor organoids, especially patient-derived organoids (PDOs) exhibit marked similarities in histopathological morphology, genomic alterations, and specific marker expression profiles to those of primary tumour tissues. They are applied in various fields including drug screening, gene editing, and identification of oncogenes. However, CAR-T therapy in the treatment of solid tumours is still at an exploratory stage. Tumour organoids offer unique advantages over other preclinical models commonly used for CAR-T therapy research, which the preservation of the biological characteristics of primary tumour tissue is critical for the study of early-stage solid tumour CAR-T therapies. Although some investigators have used this co-culture model to validate newly targeted CAR-T cells, optimise existing CAR-T cells and explore combination therapy strategies, there is still untapped potential in the co-culture models used today. This review introduces the current status of the application of tumour organoid and CAR-T cell co-culture models in recent years and commented on the limitations of the current co-cultivation model. Meanwhile, we compared the tumour organoid model with two pre-clinical models commonly used in CAR-T therapy research. Eventually, combined with the new progress of organoid technologies, optimization suggestions were proposed for the co-culture model from five perspectives: preserving or reconstructing the tumor microenvironment, systematization, vascularization, standardized culture procedures, and expanding the tumor organoids resource library, aimed at assisting related researchers to better utilize co-culture models.

Unraveling the efficacy network: A network meta-analysis of adjuvant external beam radiation therapy methods after hepatectomy.

BACKGROUND: Primary liver cancer is a malignant tumor with a high recurrence rate that significantly affects patient prognosis. Postoperative adjuvant external radiation therapy (RT) has been shown to effectively prevent recurrence after liver cancer resection. However, there are multiple RT techniques available, and the differential effects of these techniques in preventing postoperative liver cancer recurrence require further investigation. AIM: To assess the advantages and disadvantages of various adjuvant external RT methods after liver resection based on overall survival (OS) and disease-free survival (DFS) and to determine the optimal strategy. METHODS: This study involved network meta-analyses and followed the PRISMA guidelines. The data of qualified studies published before July 10, 2023, were collected from PubMed, Embase, the Web of Science, and the Cochrane Library. We included relevant studies on postoperative external beam RT after liver resection that had OS and DFS as the primary endpoints. The magnitudes of the effects were determined using risk ratios with 95% confidential intervals. The results were analyzed using R software and STATA software. RESULTS: A total of 12 studies, including 1265 patients with hepatocellular carcinoma (HCC) after liver resection, were included in this study. There was no significant heterogeneity in the direct paired comparisons, and there were no significant differences in the inclusion or exclusion criteria, intervention measures, or outcome indicators, meeting the assumptions of heterogeneity and transitivity. OS analysis revealed that patients who underwent stereotactic body radiotherapy (SBRT) after resection had longer OS than those who underwent intensity modulated radiotherapy (IMRT) or 3-dimensional conformal RT (3D-CRT). DFS analysis revealed that patients who underwent 3D-CRT after resection had the longest DFS. Patients who underwent IMRT after resection had longer OS than those who underwent 3D-CRT and longer DFS than those who underwent SBRT. CONCLUSION: HCC patients who undergo liver cancer resection must consider distinct advantages and disadvantages when choosing between SBRT and 3D-CRT. IMRT, a RT technique that is associated with longer OS than 3D-CRT and longer DFS than SBRT, may be a preferred option.

Spatiotemporal change patterns and driving factors of land surface temperature in the Yunnan-Kweichow Plateau from 2000 to 2020.

In this study, the spatiotemporal change patterns and driving factors of land surface temperature (LST) on the Yunnan-Kweichow Plateau (YKP) during 2000-2020 are investigated by using the Thermal and Reanalysis Integrating Moderate-resolution Spatial-seamless (TRIMS) LST dataset provided by National Tibetan Plateau Data Center. The YKP LST spatiotemporal change patterns are revealed at annual, seasonal, monthly, and daily scales. Furthermore, seven driving factors such as air temperature, land cover types, normalized difference vegetation index, precipitation, solar radiation, elevation, and latitude are quantified the impacts on LST spatial heterogeneity at annual scale. The main findings are as follows: (1) Annual mean LST increases by 0.016 K/year. Annual mean daytime LST slightly decreases by 0.009 K/year. Annual mean nighttime LST significantly increases by 0.042 K/year. (2) The trend and seasonal components of the daily, daily mean daytime, and daily mean nighttime LST have five and four breakpoints respectively, indicating that the variation of LST is unstable during 2000-2020 on the YKP. (3) The LST lapse rates at nighttime are generally higher than those at daytime on the YKP at the annual, seasonal, and monthly scales. The LST maximum lapse rate is 0.59 K/100 m in summer nighttime, and the LST minimum lapse rate is 0.18 K/100 m in winter daytime. (4) The controlling effects of seven factors are generally stronger in the nighttime than those in the daytime. The factors of elevation and air temperature dominate the LST spatial distribution on the YKP, with a contribution rate of >70 %. In addition, the interactions among the seven factors are all enhancing the effects on the spatial distribution of annual mean LST, including bivariate enhancement and nonlinear enhancement. This study contributes to the mitigation and adaptation to climate change of LST in the plateau and plays a theoretical reference role in formulating corresponding policies for environmental protection.

Case report: Endoscopic Hemoclip treatment for massive hematemesis caused by a Dieulafoy's lesion on duodenal papilla caused acute pancreatitis.

Background: Dieulafoy's lesion is an uncommon cause of hemorrhage of the digestive tract. It often presents with urgent and massive bleeding usually leading to shock, even death. Dieulafoy's lesions have been reported throughout the digestive tract but which occurred on duodenal papilla were particularly rare and presented challenges in the choice of hemostasis. Case presentation: A 66-year-old man with melena for 2 days was admitted. Gastrointestinal endoscopy revealed blood clots covering the duodenal papilla with oozing blood. During the procedure of trying to place a plastic stent into the duodenal papilla first, the hemorrhage began to present pulsating bleeding. The patient went into shock. With consent, two titanium clips were inserted to clamp the bleeding site to stop the bleeding. The patient complained of epigastric pain 14 h after the endoscopy. An abdominal CT scan showed signs of acute pancreatitis. Endoscopy was performed to remove the titanium clips and showed a vessel stump on the duodenal papilla. The patient was discharged from the hospital on the 14th day and followed for 6 months with no recurrence. Conclusion: This case was diagnosed with a Dieulafoy's lesion on the duodenal papilla, which has rarely been reported. Hematemesis was stopped by clamping the vessel stump with titanium clips but caused acute pancreatitis. Reviewing the treatment, electrocoagulation might be a better choice, and life support treatment, including central vena catheterization and an adequate supply of blood products, should be prepared in advance to provide extra time for the stent placement or vascular intervention treatment.

Dexmedetomidine alleviates acute lung injury by promoting Tregs differentiation via activation of AMPK/SIRT1 pathway.

OBJECTIVES: To explore the anti-inflammatory effect and the potential mechanism of dexmedetomidine in ARDS/ALI. MATERIALS AND METHODS: C57BL/6 mice and EL-4 cells were used in this research. The ALI model was established by CLP. The level of inflammatory cytokines in the lung and blood, the severity of lung injury, the expression of Foxp3, and the proportion of Tregs were detected before and after dexmedetomidine treatment. The expression of the AMPK/SIRT1 after dexmedetomidine treatment was detected in vivo and in vitro. After blocking the AMPK/SIRT1 pathway or depleting Tregs in vivo, the level of the inflammatory response, tissue injury, and Tregs differentiation were detected again to clarify the effect of dexmedetomidine. RESULTS: Dexmedetomidine significantly reduced systemic inflammation and lung injury in CLP mice. Dexmedetomidine enhanced the Foxp3 expression in the lungs and the frequency of Tregs in the spleen. Dexmedetomidine up-regulated the protein expression of p-AMPK and SIRT1 in lungs and EL-4 cells and facilitated the differentiation of naïve CD4+ T cells into Tregs in vitro. Meanwhile, DEX also increased the expression of Helios in Treg cells. CONCLUSIONS: DEX could improve ARDS/ALI by facilitating the differentiation of Tregs from naïve CD4+ T cells via activating the AMPK/SIRT1 pathway.

Suppressing CHD1L reduces the proliferation and chemoresistance in osteosarcoma.

Osteosarcoma (OS) is the most common bone malignant tumor. However, the genetic basis of OS pathogenesis is still not understood, and occurrence of chemo-resistance is a major reason for the high morbidity of OS patients. Recently, chromodomain helicase/ATPase DNA binding protein 1-like gene (CHD1L) has been identified as a gene related to malignant tumor progression. Unfortunately, its effects on OS development and drug resistance are still not understood. In the study, we attempted to investigate the effects of CHD1L on tumorigenesis and chemoresistance in OS. We found that CHD1L expression was markedly up-regulated in OS samples, especially in cisplatin (cDDP)-resistant patients. We also showed that OS cells with CHD1L knockdown were more sensitive to cDDP treatment with lower IC50 values. In addition, we found that CHD1L deletion markedly reduced cell proliferation and induced apoptosis in OS cells with cDDP resistance. Moreover, the properties of cancer stem cells were highly suppressed in cDDP-resistant OS cells following CHD1L knockdown. Furthermore, multidrug resistance protein 1 (MDR-1) expression levels were dramatically decreased in OS cells with cDDP resistance when CHD1L was suppressed. Functional analysis indicated that CHD1L knockdown clearly restrained the activation of ERK1/2, protein kinase B (AKT) and NF-κB signaling pathways in cDDP-resistant OS cells. Consistently, animal experiments suggested that CHD1L suppression mitigated cDDP resistance in the generated in vivo xenografts. Collectively, CHD1L could modulate chemoresistance of OS cells to cDDP, and thus may be inspiring findings for overcoming drug resistance in OS.

Regulation of the expression of proinflammatory cytokines induced by SARS-CoV-2.

An outbreak of a novel coronavirus was reported in Wuhan, China, in late 2019. It has spread rapidly through China and many other countries, causing a global pandemic. Since February 2020, over 28 countries/regions have reported confirmed cases. Individuals with the infection known as coronavirus disease-19 (COVID-19) have similar clinical features as severe acute respiratory syndrome first encountered 17 years ago, with fever, cough, and upper airway congestion, along with high production of proinflammatory cytokines (PICs), which form a cytokine storm. PICs induced by COVID-19 include interleukin (IL)-6, IL-17, and monocyte chemoattractant protein-1. The production of cytokines is regulated by activated nuclear factor-kB and involves downstream pathways such as Janus kinase/signal transducers and activators transcription. Protein expression is also regulated by post-translational modification of chromosomal markers. Lysine residues in the peptide tails stretching out from the core of histones bind the sequence upstream of the coding portion of genomic DNA. Covalent modification, particularly methylation, activates or represses gene transcription. PICs have been reported to be induced by histone modification and stimulate exudation of hyaluronic acid, which is implicated in the occurrence of COVID-19. These findings indicate the impact of the expression of PICs on the pathogenesis and therapeutic targeting of COVID-19.

Massive gastrointestinal bleeding caused by a Dieulafoy's lesion in a duodenal diverticulum: A case report.

BACKGROUND: Dieulafoy's lesion is a rare vascular abnormality characterized by a small abnormally dilated artery that runs a tortuous course in the submucosa. There is usually no ulcer present in Dieulafoy's lesions and the overlying mucosa is most often normal. Bleeding caused by a Dieulafoy's lesion is usually urgent, massive, life-threatening and prone to recurrence. Dieulafoy's lesions have been reported throughout the digestive tract although the majority of them have been found in the upper digestive tract especially the stomach and duodenum. However, a Dieulafoy's lesion occurring inside a duodenal diverticulum is very rare. CASE SUMMARY: A 74-year-old Asian male with epigastric pain, hematemesis and melena was admitted to our clinic. Before admission, the patient had vomited 500 mL of dark red blood, and passed 200 g of black tarry stool. Conservative management was first undertaken as the patient had not been fasting. However, hemorrhage recurred and the patient went into shock. Urgent endoscopy was performed and a diverticulum of 1.8 cm × 1.2 cm × 0.8 cm was found on the anterior wall of the descending duodenum. The diverticulum was covered with a blood clot. After the clot was removed, an artery stump was observed in the diverticulum with a diameter of 2-3 mm. Two titanium hemostatic clips were inserted to clamp the vessel stump. The patient was discharged 7 d post-endoscopy and followed for 6 mo with no recurrence. CONCLUSION: This case was diagnosed with a Dieulafoy's lesion inside a duodenal diverticulum which has rarely been reported. Hematemesis was stopped by clamping the vessel stump with titanium clips. No complications occurred.

Nanomaterials: Applications in the diagnosis and treatment of pancreatic cancer.

Pancreatic cancer (PC) remains one of the leading causes of cancer-related death in human sowing to missed early and effective diagnosis. The inability to translate research into clinical trials and to target chemotherapy drugs to tumors is a major obstacle in PC treatment. Compared with traditional cancer detection methods, the method combining existing clinical diagnosis and detection systems with nanoscale components using novel nanomaterials shows higher sensitivity and specificity. Nanomaterials can interact with biological systems to efficiently and accurately detect and monitor biological events during diagnosis and treatment. With the advance of experimental and engineering technology, more nanomaterials will begin the transition to clinical trials for their validation. This paper describes a number of nanomaterials used in the diagnosis and treatment of PC.

LAMB3 mediates apoptotic, proliferative, invasive, and metastatic behaviors in pancreatic cancer by regulating the PI3K/Akt signaling pathway.

The poor prognosis of patients with pancreatic ductal adenocarcinoma (PDAC) is partially attributed to the invasive and metastatic behavior of this disease. Laminin subunit beta-3 (LAMB3) encodes one of the three subunits of LM-332, an extracellular matrix protein secreted by cultured human keratinocytes. In addition, LAMB3 is involved in the invasive and metastatic abilities of some types of cancer, including colon, pancreas, lung, cervix, stomach, and prostate cancer, but the role and mechanism of LAMB3 in PDAC have not been previously determined. Herein, we tentatively investigated the role of LAMB3 in the malignant biological behavior of PDAC. In this study, we demonstrated that LAMB3 is upregulated in PDAC. Inhibition of LAMB3 abrogated the tumorigenic outcomes of PI3K/Akt signaling pathway activation, including those involving cell cycle arrest, cell apoptosis, proliferation, invasion and migration in vitro, and tumor growth and liver metastasis in vivo. Our results showed that LAMB3 could mediate cell cycle arrest and apoptosis in PDAC cells and alter the proliferative, invasive, and metastatic behaviors of PDAC by regulating the PI3K/Akt signaling pathway. LAMB3 may be a novel therapeutic target for the treatment of PDAC in the future.

Effect of lentiviral vector-mediated overexpression of hypoxia-inducible factor 1 alpha delivered by pluronic F-127 hydrogel on brachial plexus avulsion in rats.

Brachial plexus avulsion often results in massive motor neuron death and severe functional deficits of target muscles. However, no satisfactory treatment is currently available. Hypoxia-inducible factor 1α is a critical molecule targeting several genes associated with ischemia-hypoxia damage and angiogenesis. In this study, a rat model of brachial plexus avulsion-reimplantation was established, in which C5-7 ventral nerve roots were avulsed and only the C6 root reimplanted. Different implants were immediately injected using a microsyringe into the avulsion-reimplantation site of the C6 root post-brachial plexus avulsion. Rats were randomly divided into five groups: phosphate-buffered saline, negative control of lentivirus, hypoxia-inducible factor 1α (hypoxia-inducible factor 1α overexpression lentivirus), gel (pluronic F-127 hydrogel), and gel + hypoxia-inducible factor 1α (pluronic F-127 hydrogel + hypoxia-inducible factor 1α overexpression lentivirus). The Terzis grooming test was performed to assess recovery of motor function. Scores were higher in the hypoxia-inducible factor 1α and gel + hypoxia-inducible factor 1α groups (in particular the gel + hypoxia-inducible factor 1α group) compared with the phosphate-buffered saline group. Electrophysiology, fluorogold retrograde tracing, and immunofluorescent staining were further performed to investigate neural pathway reconstruction and changes of neurons, motor endplates, and angiogenesis. Compared with the phosphate-buffered saline group, action potential latency of musculocutaneous nerves was markedly shortened in the hypoxia-inducible factor 1α and gel + hypoxia-inducible factor 1α groups. Meanwhile, the number of fluorogold-positive cells and ChAT-positive neurons, neovascular area (labeled by CD31 around avulsed sites in ipsilateral spinal cord segments), and the number of motor endplates in biceps brachii (identified by α-bungarotoxin) were all visibly increased, as well as the morphology of motor endplate in biceps brachil was clear in the hypoxia-inducible factor 1α and gel + hypoxia-inducible factor 1α groups. Taken together, delivery of hypoxia-inducible factor 1α overexpression lentiviral vectors mediated by pluronic F-127 effectively promotes spinal root regeneration and functional recovery post-brachial plexus avulsion. All animal procedures were approved by the Institutional Animal Care and Use Committee of Guangdong Medical University, China.

ZBP-89 and Sp1 contribute to Bak expression in hepatocellular carcinoma cells.

BACKGROUND: Kruppel family member zinc binding protein 89 (ZBP-89), also known as ZNF148, regulates Bak expression via binding to GC-rich promoter domain. It is not clear if other GC-rich binding factors, such as Sp family members, can interact with ZBPp-89 on Bak expression. This study aims to elucidate the mechanism of Bak expression regulation by ZBP-89 and Sp proteins, based on in vitro experiment and The Cancer Genome Atlas (TCGA) hepatocellular carcinoma (HCC) data cohort. METHODS: We downloaded TCGA hepatocellular carcinoma (HCC) cohort data to analysis the association of Bak transcription level with ZBP-89 and Sp proteins transcription level. HCC cell lines and liver immortal non-tumour cell lines were used for mechanism study, including western blotting analysis, expression vector mediated gene expression and siRNA interference. RESULTS: Results showed that cancer tissues have higher Bak transcription level compared with adjacent non-cancer tissues. Bak transcription level was correlated with Sp1 and Sp3 expression level, while no correlation was found in ZBP-89 and Bak, neither Sp2 nor Sp4. Mithramycin A (MMA) induced Bak expression in a dose-dependent manner. Western blotting results showed Sp1 overexpression increased Bak expression both in liver immortal non-tumour cells and HCC cells. Interference Sp1 expression could inhibit Bak expression alone. ZBP-89 siRNA suppressed Bak expression even in the presence of MMA treatment and S1 overexpression. Additionally, Bak and Sp1 level were associated with HCC patient survival. CONCLUSIONS: Bak expression required ZBP-89 and Sp1 cooperative regulation simultaneously.

Epigallocatechin-3-gallate Modulates MicroRNA Expression Profiles in Human Nasopharyngeal Carcinoma CNE2 Cells.

BACKGROUND: Epigallocatechin-3-gallate (EGCG) has exhibited antitumor properties in several types of cancers, including nasopharyngeal carcinoma (NPC), but the molecular mechanisms underlying this function remain incompletely understood. The aim of the present study was to characterize the global impact of EGCG on the expression of microRNAs (miRNAs) in NPC cells. METHODS: Using microarray analysis, the alterations of miRNA expression profiles were investigated in EGCG-treated CNE2 cells. Furthermore, the target genes and signaling pathways regulated by EGCG-specific miRNAs were identified using target prediction program and gene ontology analysis. RESULTS: A total of 14 miRNAs exhibited >2-fold expression changes in a dose-dependent manner after treatment with 20 µmol/L and 40 µmol/L EGCG. Totally 43, 49, and 52 target genes from these differentially expressed miRNAs were associated with the apoptosis, cell cycle regulation, and cell proliferation, respectively. A total of 66 signaling pathways, primarily involved in cancer development and lipid and glucose metabolism, were shown to be regulated by EGCG-specific miRNAs. CONCLUSION: EGCG induces considerable alterations of miRNA expression profiles in CNE2 cells, which provides mechanistic insights into cellular responses and antitumor activity mediated by EGCG.

Expression and clinical significance of C14orf166 in esophageal squamous cell carcinoma.

C14orf166, a 28 kD protein regulating RNA transcription and translation, may serve a critical role in oncogenesis. The aim of the current study was to explore the association between C14orf166 expression and esophageal squamous cell carcinoma (ESCC) and to draw attention to the association between C14orf166 and the initiation, progression and prognosis of ESCC. C14orf166 expression in ESCC and paired normal tissues was detected by immunohistochemical staining, western blotting and reverse transcription­quantitative polymerase chain reaction, and the association between C14orf166 expression and clinicopathological characters of ESCC was analyzed. Survival analysis was used to assess the prognostic significance of C14orf166 and it was observed that C14orf166 expression was higher in the ESCC tissues when compared with adjacent non­cancerous tissues at protein (P<0.001) and mRNA levels (P<0.001). There was a significant difference in T stage, lymph node metastasis and TNM stage in patients categorized according to different C14orf166 expression levels. The overexpression of C14orf166 was associated with a shorter overall survival and disease­free survival, and multivariate analysis indicated that C14orf166 was an independent prognostic indicator. The present study indicates that the expression of C14orf166 is elevated in ESCC, and is potentially a valuable prognostic predictor for ESCC.

Evolution of mixing width induced by general Rayleigh-Taylor instability.

Turbulent mixing induced by Rayleigh-Taylor (RT) instability occurs ubiquitously in many natural phenomena and engineering applications. As the simplest and primary descriptor of the mixing process, the evolution of mixing width of the mixing zone plays a notable role in the flows. The flows generally involve complex varying acceleration histories and widely varying density ratios, two dominant factors affecting the evolution of mixing width. However, no satisfactory theory for predicting the evolution has yet been established. Here a theory determining the evolution of mixing width in general RT flows is established to reproduce, first, all of the documented experiments conducted for diverse (i.e., constant, impulsive, oscillating, decreasing, increasing, and complex) acceleration histories and all density ratios. The theory is established in terms of the conservation principle, with special consideration given to the asymmetry of the volume-averaged density fields occurring in actual flows. The results reveal the sensitivity or insensitivity of the evolution of a mixing front of a neighboring light or heavy fluid to the degree of asymmetry and thus explain the distinct evolutions in two experiments with the same configurations.

Nemo-like kinase as a negative regulator of nuclear receptor Nurr1 gene transcription in prostate cancer.

BACKGROUND: Nurr1, a member of the orphan receptor family, plays an important role in several types of cancer. Our previous work demonstrated that increased expression of Nurr1 plays a significant role in the initiation and progression of prostate cancer (PCa), though the mechanisms for regulation of Nurr1 expression remain unknown. In this study, we investigated the hypothesis that Nemo-like kinase (NLK) is a key regulator of Nurr1 expression in PCa. METHODS: Immunohistochemistry and Western blot analysis were used to evaluate levels of NLK and Nurr1 in prostatic tissues and cell lines. The effects of overexpression or knockdown of Nurr1 were evaluated in PCa cells through use of PCR, Western blots and promoter reporter assays. The role of Nurr1 promoter cis element was studied by creation of two mutant Nurr1 promoter luciferase constructs, one with a mutated NF-κB binding site and one with a mutated CREB binding site. In addition, three specific inhibitors were used to investigate the roles of these proteins in transcriptional activation of Nurr1, including BAY 11-7082 (NF-κB inhibitor), KG-501 (CREB inhibitor) and ICG-001 (CREB binding protein, CBP, inhibitor). The function of CBP in NLK-mediated regulation of Nurr1 expression was investigated using immunofluorescence, co-immunoprecipitation (Co-IP) and chromatin immunoprecipitation assays (ChIPs). RESULTS: NLK expression was inversely correlated with Nurr1 expression in prostate cancer tissues and cell lines. Overexpression of NLK suppressed Nurr1 promoter activity, leading to downregulation of Nurr1 expression. In contrast, knockdown of NLK demonstrated opposite results, leading to upregulation of Nurr1. When compared with the wild-type Nurr1 promoter, mutation of NF-κB- and CREB-binding sites of the Nurr1 promoter region significantly reduced the upregulation of Nurr1 induced by knockdown of NLK in LNCaP cells; treatment with inhibitors of CREB, CBP and NF-κB led to similar results. We also found that NLK directly interacts with CBP, that knockdown of NLK significantly increases the recruitment of CBP to both NF-κB- and CREB-binding sites, and that regulation of NLK on Nurr1 expression is abrogated by knockdown of CBP. CONCLUSIONS: Our results suggest that NLK inhibits transcriptional activation of Nurr1 gene by impeding CBP's role as a co-activator of NF-κB and CREB in prostate cancer.

Huangqin-Tang Ameliorates TNBS-Induced Colitis by Regulating Effector and Regulatory CD4(+) T Cells.

Huangqin-Tang decoction (HQT) is a classic traditional Chinese herbal formulation that is widely used to ameliorate the symptoms of gastrointestinal disorders, including inflammatory bowel disease (IBD). This study was designed to investigate the therapeutic potential and immunological regulatory activity of HQT in experimental colitis in rats. Using an animal model of colitis by intrarectally administering 2,4,6-trinitrobenzenesulfonic acid (TNBS), we found that administration of HQT significantly inhibited the severity of TNBS-induced colitis in a dose-dependent manner. In addition, treatment with HQT produced better results than that with mesalazine, as shown by improvedweight loss bleeding and diarrhoea scores, colon length, and intestinal inflammation. As for potential immunological regulation of HQT action, the percentages of Th1 and Th17 cells were reduced, but those Th2 and Treg cells were enhanced in LPMCs after HQT treatment. Additionally, HQT lowered the levels of Th1/Th17-associated cytokines but increased production of Th2/Treg-associated cytokines in the colon and MLNs. Furthermore, we observed a remarkable suppression of the Th1/Th17-associated transcription factors T-bet and ROR-γt. However, expression levels of the Th2/Treg-associated transcription factors GATA-3 and Foxp3 were enhanced during treatment with HQT. Our results suggest that HQT has the therapeutic potential to ameliorate TNBS-induced colitis symptoms. This protective effect is possibly mediated by its effects on CD4(+) T cells subsets.

[p38 signal pathway regulates osteogenic differentiation of maxillary primordium mesenchymal cells].

PURPOSE: To explore whether p38 signal pathway regulates osteogenic differentiation of maxillary primordium mesenchymal cells. METHODS: The first passage of maxillary primordium mesenchymal cells (MPMCs) from E12.5 embryos were cultured in the osteogenic medium, and 10 nM SB203580 (an inhibitor of phosphorylation of p38) was added in the medium in the experimental group for 1 week. Then immunofluorescence staining was applied to detect the phosphorylation of p38 in MPMCs. Brdu label and immunofluorescence staining were used to detect the proliferation of MPMCs. ALP staining and qPCR were used to detect the mRNA expression of ALP, Runx2, OCN and OPN in MPMCs. ALP staining and PCR were used to evaluate the osteogenic capability of MPMCs. SPSS 18.0 software package was used to analyze the data. RESULTS: Osteogenic induction could promote phosphorylation of p38, inhibit phosphorylation of p38 and proliferation of MPMCs, down-regulate the expression of ALP, Runx2, OCN and OPN, thus weaken the ALP staining in MPMCs. CONCLUSIONS: p38 signal pathway regulates osteogenic differentiation of MPMCs in vitro.

Baicalin attenuates TNBS-induced colitis in rats by modulating the Th17/Treg paradigm.

Baicalin, a flavonoid, has a wide range of pharmacological properties, including immunomodulation. The objective of this study was to investigate the effect of baicalin on the balance of T helper 17 (Th17) and regulatory T (Treg) cells in a colitis model. The rat colitis model was induced by 2,4,6-trinitrobenzene sulfonic acid (TNBS). Baicalin (10 ml/kg, each) or mesalazine (positive control) was then administered orally for 7 days. Inflammatory and immunological responses were evaluated by pathology, enzyme-linked immunosorbent assay, real-time polymerase chain reaction, western blot analysis, and flow cytometry. Our study showed that baicalin not only significantly attenuated TNBS-induced colitis by reducing the disease activity index as well as macroscopic and microscopic scores, but it also improved the weight loss and shortening of the colon. Baicalin treatment also induced a significant decrease in the levels of inflammatory mediators, including the myeloperoxidase activity, the levels of tumor necrosis factor α, IL-1ß, and Th1-related cytokines IL-12 and IFN-γ. Furthermore, the beneficial effects of baicalin seem to be associated with regulation of the Th17 and Treg paradigm. We found that administration of baicalin significantly downregulated the number of Th17 cells and the levels of Th17-related cytokines (IL-17 and IL-6) and retinoic acid receptor-related orphan receptor γt. In contrast, there was an increase in Treg cells numbers, Treg-related cytokines transforming growth factor-ß and IL-10, and forkhead box P3. Our results suggest that the anti-inflammatory effect of baicalin may be linked to modulation of the balance between Th17 and Treg cells in TNBS-induced ulcerative colitis.