ZBTB46 coordinates angiogenesis and immunity to control tumor outcome.

Tumor angiogenesis and immunity show an inverse correlation in cancer progression and outcome1. Here, we report that ZBTB46, a repressive transcription factor and a widely accepted marker for classical dendritic cells (DCs)2,3, controls both tumor angiogenesis and immunity. Zbtb46 was downregulated in both DCs and endothelial cells by tumor-derived factors to facilitate robust tumor growth. Zbtb46 downregulation led to a hallmark pro-tumor microenvironment (TME), including dysfunctional vasculature and immunosuppressive conditions. Analysis of human cancer data revealed a similar association of low ZBTB46 expression with an immunosuppressive TME and a worse prognosis. In contrast, enforced Zbtb46 expression led to TME changes to restrict tumor growth. Mechanistically, Zbtb46-deficient endothelial cells were highly angiogenic, and Zbtb46-deficient bone marrow progenitors upregulated Cebpb and diverted the DC program to immunosuppressive myeloid lineage output, potentially explaining the myeloid lineage skewing phenomenon in cancer4. Conversely, enforced Zbtb46 expression normalized tumor vessels and, by suppressing Cebpb, skewed bone marrow precursors toward immunostimulatory myeloid lineage output, leading to an immune-hot TME. Remarkably, Zbtb46 mRNA treatment synergized with anti-PD1 immunotherapy to improve tumor management in preclinical models. These findings identify ZBTB46 as a critical factor for angiogenesis and for myeloid lineage skewing in cancer and suggest that maintaining its expression could have therapeutic benefits.

Genetic analysis of a child with severe intellectual disability caused by a novel variant in the FERM domain of the FRMPD4 protein.

Intellectual developmental disorder, X-linked 104 (XLID104), caused by the FRMPD4 gene variant, is a rare X-linked genetic disease that primarily manifests as intellectual disability (ID) and language delay, and may be accompanied by behavioural abnormalities. Currently, only 11 patients from four families have been reported to carry FRMPD4 gene variants. Here, we report a rare case of a Chinese patient with XLID104 who was presented with severe ID and language impairment. Genetic testing results showed that the patient had a novel hemizygous variant on FRMPD4 inherited from the heterozygous variant NM_001368397: c.1772A>C (p.Glu591Ala) carried by his mother. To our knowledge, this variant has not been reported previously. Western blot results for the recombinant plasmid constructed in vitro indicated that the expression of the mutant protein may be reduced. Using molecular dynamics simulations, we predicted that the mutant protein may affect the interaction of the FRMPD4 protein with DLG4. In this study, we expand the spectrum of FRMPD4 variants and suggest that the clinical awareness of the genetic diagnosis of nonsyndromic ID should be strengthened.

Clinical characteristics of Guillain-Barré syndrome in patients with primary Sjögren's syndrome.

To investigate the clinical characteristics of Guillain-Barré syndrome (GBS) in patients with primary Sjögren's syndrome (SS). Records of patients with positive anti-SSA antibodies hospitalized in the Beijing Tiantan Hospital between December 2011 and May 2020 were retrieved. Patients who fulfilled the criteria for diagnosis of GBS and primary SS were included, and their clinical data were analyzed. Among the 785 patients with positive anti-SSA, 52 patients were identified in this study. They were 27 males and 25 females with median age of 59 years old. Besides anti-SSA antibodies, multiple autoantibodies were detected in these patients including antinuclear antibody, anti-Ro52, anti-mitochondrial M2, anti-thyroid peroxidase and anti-thyroglobulin autoantibodies. Preceding infection was reported in 42 patients. Hyporeflexia/areflexia and limbs weakness were the most common manifestation and 35 patients presented cranial nerve injuries. GBS disability score of 3, 4 and 5 was scaled in 28 (53.8%), 15 (28.8%) and 3 (5.8%) patients respectively. Forty-six patients received intravenous immunoglobulin (IVIG) monotherapy, 5 patients were treated by IVIG plus glucocorticoids, and 51 patients improved during hospitalization. The frequency of male gender among the patients with both GBS and primary SS suggests an independent onset of GBS and the co-existence of these autoimmune diseases in patients with multiple autoantibodies. Majority of patients with GBS and primary SS experience benign disease course.

Wiskott-Aldrich syndrome: A new synonym mutation in the WAS gene.

Wiskott-Aldrich syndrome (WAS) is a rare X-linked recessive primary immunodeficiency disorder. Mutations in the WAS gene are considered to be the primary cause of WAS. In this work, we report a boy who presented with intracranial hemorrhage (ICH) as an initial symptom and detects a novel pathogenic synonymous mutation in his WAS gene. His mother was a carrier of the mutant gene. The mutation, located at position c.273 (c.273 G>A) in exon 2, is a synonym mutation and predicted to affect protein expression by disrupting gene splicing. This study summarizes the diagnosis and treatment process of the patient and expands the genetic spectrum of WAS.

Dystonia, spastic tetraplegia, and ataxia due to a novel mutation in the dynamin domain of OPA1.

Movement disorders manifest in various hereditary neurodegenerative diseases. We reported a young man who presented with progressive upper limb dystonia, spastic tetraplegia, and ataxia. Whole-exome sequencing (WES) revealed a novel variant, c.2357A > G, in the dynamin domain of OPA1. No mtDNA deletion was detected in muscle by long-range PCR. Atrophy and decreased glucose metabolism of the basal ganglia were discovered. Decreased mtDNA copy number, fragmented mitochondria, slightly impaired oxidative phosphorylation, and increased autophagy were detected in mutant fibroblasts. Evident oxidative phosphorylation impairment and mtDNA deletions were not involved in the pathogenicity of this mutation unlike mutations in the GTPase domain of OPA1.

Systemic delivery of murine SOD2 mRNA to experimental abdominal aortic aneurysm mitigates expansion and rupture.

Background: Oxidative stress is implicated in the pathogenesis and progression of abdominal aortic aneurysm (AAA). Antioxidant delivery as a therapeutic for AAA is of substantial interest although clinical translation of antioxidant therapy has met with significant challenges due to limitations in achieving sufficient antioxidant levels at the site of AAA. We posit that nanoparticle-based approaches hold promise to overcome challenges associated with systemic administration of antioxidants. Methods: We employed a peptide-based nanoplatform to overexpress a key modulator of oxidative stress, superoxide dismutase 2 (SOD2). The efficacy of systemic delivery of SOD2 mRNA as a nanotherapeutic agent was studied in two different murine AAA models. Unbiased mass spectrometry-enabled proteomics and high-dimensional bioinformatics were used to examine pathways modulated by SOD2 overexpression. Results: The murine SOD2 mRNA sequence was mixed with p5RHH, an amphipathic peptide capable of delivering nucleic acids in vivo to form self-assembled nanoparticles of ∼55 nm in diameter. We further demonstrated that the nanoparticle was stable and functional up to four weeks following self-assembly when coated with hyaluronic acid. Delivery of SOD2 mRNA mitigated the expansion of small AAA and largely prevented rupture. Mitigation of AAA was accompanied by enhanced SOD2 protein expression in aortic wall tissue. Concomitant suppression of nitric oxide, inducible nitric oxide synthase expression, and cell death was observed. Proteomic profiling of AAA tissues suggests that SOD2 overexpression augments levels of microRNAs that regulate vascular inflammation and cell apoptosis, inhibits platelet activation/aggregation, and downregulates mitogen-activated protein kinase signaling. Gene set enrichment analysis shows that SOD2 mRNA delivery is associated with activation of oxidative phosphorylation, lipid metabolism, respiratory electron transportation, and tricarboxylic acid cycle pathways. Conclusions: These results confirm that SOD2 is key modulator of oxidative stress in AAA. This nanotherapeutic mRNA delivery approach may find translational application in the medical management of small AAA and the prevention of AAA rupture.

Synergizing Deep Learning-Enabled Preprocessing and Human-AI Integration for Efficient Automatic Ground Truth Generation.

The progress of incorporating deep learning in the field of medical image interpretation has been greatly hindered due to the tremendous cost and time associated with generating ground truth for supervised machine learning, alongside concerns about the inconsistent quality of images acquired. Active learning offers a potential solution to these problems of expanding dataset ground truth by algorithmically choosing the most informative samples for ground truth labeling. Still, this effort incurs the costs of human labeling, which needs minimization. Furthermore, automatic labeling approaches employing active learning often exhibit overfitting tendencies while selecting samples closely aligned with the training set distribution and excluding out-of-distribution samples, which could potentially improve the model's effectiveness. We propose that the majority of out-of-distribution instances can be attributed to inconsistent cross images. Since the FDA approved the first whole-slide image system for medical diagnosis in 2017, whole-slide images have provided enriched critical information to advance the field of automated histopathology. Here, we exemplify the benefits of a novel deep learning strategy that utilizes high-resolution whole-slide microscopic images. We quantitatively assess and visually highlight the inconsistencies within the whole-slide image dataset employed in this study. Accordingly, we introduce a deep learning-based preprocessing algorithm designed to normalize unknown samples to the training set distribution, effectively mitigating the overfitting issue. Consequently, our approach significantly increases the amount of automatic region-of-interest ground truth labeling on high-resolution whole-slide images using active deep learning. We accept 92% of the automatic labels generated for our unlabeled data cohort, expanding the labeled dataset by 845%. Additionally, we demonstrate expert time savings of 96% relative to manual expert ground-truth labeling.

Molecular and immunological characteristics of postoperative relapse in lymph node-positive esophageal squamous cell cancer.

BACKGROUND: The molecular and immunological characteristics of primary tumors and positive lymph nodes in esophageal squamous cell carcinoma (ESCC) are unknown and the relationship with recurrence is unclear, which this study attempted to explore. METHODS: A total of 30 ESCC patients with lymph node positive (IIB-IVA) were enrolled. Among them, primary tumor and lymph node specimens were collected from each patient, and subjected to 551-tumor-targeted DNA sequencing and 289-immuno-oncology RNA panel sequencing to identify the different molecular basis and immunological features, respectively. RESULTS: The primary tumors exhibited a higher mutation burden than lymph nodes (p < 0.001). One-year recurrent ESCC exhibited a higher Mucin16 (MUC16) mutation rate (p = 0.038), as well as univariate and multivariate analysis revealed that MUC16 mutation is independent genetic factor associated with reduced relapse-free survival (univariate, HR: 5.39, 95% CI: 1.67-17.4, p = 0.005; multivariate, HR: 7.36, 95% CI: 1.79-30.23, p = 0.006). Transcriptomic results showed non-relapse group had higher cytolytic activity (CYT) score (p = 0.025), and was enriched in the IFN-α pathway (p = 0.036), while those in the relapsed group were enriched in the TNF-α/NF-κB (p = 0.001) and PI3K/Akt pathway (p = 0.014). CONCLUSION: The difference in molecular characteristics between primary lesions and lymph nodes may be the cause of the inconsistent clinical outcomes. Mutations of MUC16 and poor immune infiltration are associated with rapid relapse of nodes-positive ESCC.

Curcumin for gastric cancer: Mechanism prediction via network pharmacology, docking, and in vitro experiments.

BACKGROUND: Curcumin originates from the natural herb turmeric, and its antitumor effects have been known about for a long time. However, the mechanism by which curcumin affects gastric cancer (GC) has not been elucidated. AIM: To elucidate the potential mechanisms of curcumin in the treatment of GC. METHODS: Network pharmacological approaches were used to perform network analysis of Curcumin. We first analyzed Lipinski's Rule of Five for the use of Curcumin. Curcumin latent targets were predicted using the PharmMapper, SwissTargetPrediction and DrugBank network databases. GC disease targets were mined through the GeneCard, OMIM, DrugBank and TTD network databases. Then, GO enrichment, KEGG enrichment, protein-protein interaction (PPI), and overall survival analyses were performed. The results were further verified through molecular docking, differential expression analysis and cell experiments. RESULTS: We identified a total of 48 curcumin-related genes with 31 overlapping GC-related targets. The intersection targets between curcumin and GC have been enriched in 81 GO biological processes and 22 significant pathways. Following PPI analysis, 6 hub targets were identified, namely, estrogen receptor 1 (ESR1), epidermal growth factor receptor (EGFR), cytochrome P450 family 3 subfamily A member 4 (CYP3A4), mitogen-activated protein kinase 14 (MAPK14), cytochrome P450 family 1 subfamily A member 2 (CYP1A2), and cytochrome p450 family 2 subfamily B member 6 (CYP2B6). These factors are correlated with decreased survival rates among patients diagnosed with GC. Molecular docking analysis further substantiated the strong binding interactions between Curcumin and the hub target genes. The experimental findings demonstrated that curcumin not only effectively inhibits the growth of BGC-823 cells but also suppresses their proliferation. mRNA levels of hub targets CYP3A4, MAPK14, CYP1A2, and CYP2B6 in BGC-823 cells were significantly increased in each dose group. CONCLUSION: Curcumin can play an anti-GC role through a variety of targets, pathways and biological processes.

Glycolysis inhibition induces anti-tumor central memory CD8+T cell differentiation upon combination with microwave ablation therapy.

Minimally invasive thermal therapy is a successful alternative treatment to surgery in solid tumors with high complete ablation rates, however, tumor recurrence remains a concern. Central memory CD8+ T cells (TCM) play important roles in protection from chronic infection and cancer. Here we find, by single-cell RNA analysis of human breast cancer samples, that although the memory phenotype of peripheral CD8+ T cells increases slightly after microwave ablation (MWA), the metabolism of peripheral CD8+ T cells remains unfavorable for memory phenotype. In mouse models, glycolysis inhibition by 2-deoxy-D-glucose (2DG) in combination with MWA results in long-term anti-tumor effect via enhancing differentiation of tumor-specific CD44hiCD62L+CD8+ TCM cells. Enhancement of CD8+ TCM cell differentiation determined by Stat-1, is dependent on the tumor-draining lymph nodes (TDLN) but takes place in peripheral blood, with metabolic remodeling of CD8+ T cells lasting the entire course of the the combination therapy. Importantly, in-vitro glycolysis inhibition in peripheral CD8+ T cells of patients with breast or liver tumors having been treated with MWA thrice leads to their differentiation into CD8+ TCM cells. Our work thus offers a potential strategy to avoid tumor recurrence following MWA therapy and lays down the proof-of-principle for future clinical trials.

Weiwei Decoction alleviates gastric intestinal metaplasia through the olfactomedin 4/nucleotide-binding oligomerization domain 1/caudal-type homeobox gene 2 signaling pathway.

BACKGROUND: Gastric intestinal metaplasia (IM) is a precancerous lesion that is associated with an elevated risk of gastric carcinogenesis. Weiwei Decoction (WWD) is a promising traditional Chinese herbal formula widely employed in clinical for treating IM. Previous studies suggested the potential involvement of the olfactomedin 4 (OLFM4)/nucleotide-binding oligomerization domain 1 (NOD1)/caudal-type homeobox gene 2 (CDX2) signaling pathway in IM regulation. AIM: To verify the regulation of the OLFM4/NOD1/CDX2 pathway in IM, specifically investigating WWD's effectiveness on IM through this pathway. METHODS: Immunohistochemistry for OLFM4, NOD1, and CDX2 was conducted on tissue microarray. GES-1 cells treated with chenodeoxycholic acid were utilized as IM cell models. OLFM4 short hairpin RNA (shRNA), NOD1 shRNA, and OLFM4 pcDNA were transfected to clarify the pathway regulatory relationships. Protein interactions were validated by co-immunoprecipitation. To explore WWD's pharmacological actions, IM rat models were induced using N-methyl-N'-nitro-N-nitrosoguanidine followed by WWD gavage. Gastric cells were treated with WWD-medicated serum. Cytokines and chemokines content were assessed by enzyme-linked immunosorbent assay and quantitative reverse transcription polymerase chain reaction. RESULTS: The OLFM4/NOD1/CDX2 axis was a characteristic of IM. OLFM4 exhibited direct binding and subsequent down-regulation of NOD1, thereby sustaining the activation of CDX2 and promoting the progression of IM. WWD improved gastric mucosal histological lesions while suppressing intestinal markers KLF transcription factor 4, villin 1, and MUCIN 2 expression in IM rats. Regarding pharmacological actions, WWD suppressed OLFM4 and restored NOD1 expression, consequently reducing CDX2 at the mRNA and protein levels in IM rats. Parallel regulatory mechanisms were observed at the protein level in IM cells treated with WWD-medicated serum. Furthermore, WWD-medicated serum treatment strengthened OLFM4 and NOD1 interaction. In case of anti-inflammatory, WWD restrained interleukin (IL)-6, interferon-gamma, IL-17, macrophage chemoattractant protein-1, macrophage inflammatory protein 1 alpha content in IM rat serum. WWD-medicated serum inhibited tumor necrosis factor alpha, IL-6, IL-8 transcriptions in IM cells. CONCLUSION: The OLFM4/NOD1/CDX2 pathway is involved in the regulation of IM. WWD exerts its therapeutic efficacy on IM through the pathway, additionally attenuating the inflammatory response.

Bile Acids Promote Hepatic Biotransformation and Excretion of Aflatoxin B1 in Broiler Chickens.

Aflatoxin B1 (AFB1) is a hazardous mycotoxin that often contaminates animal feed and may potentially induce severe liver damage if ingested. The liver is the primary organ responsible for AFB1 detoxification through enzyme-catalyzed xenobiotic metabolism and bile acid (BA)-associated excretion. In this study, we sought to investigate whether exogenous BA improves hepatic AFB1 detoxification to alleviate AFB1-induced liver injury in broiler chickens. Five-day-old broiler chicks were randomly assigned to three groups. CON and AFB1 received a basal diet; AFB1 + BA received a basal diet with 250 mg/kg BA for 20 days. After a 3-day pre-feed, AFB1 and AFB1 + BA were daily gavaged with 250 µg/kg BW AFB1, while CON received gavage solvent for AFB1 treatment. Dietary BA supplementation protected chickens from AFB1-induced hepatic inflammation and oxidative stress. The hepatic biotransformation of AFB1 to its metabolite AFBO was improved, with accelerated excretion to the gallbladder and cecum. Accordantly, AFB1-induced down-regulation of detoxification genes, including cytochrome P450 enzymes, glutathione S-transferases, and the bile salt export pump, was rescued by BA supplementation. Moreover, liver X receptor α, suppressed by AFB1, was enhanced in BA-treated broiler chickens. These results indicate that dietary BA supplementation improves hepatic AFB1 detoxification and excretion through LXRα-involved regulation of xenobiotic enzymes.

Effects of total flavonoids of Hippophae rhamnoides L. on intracellular free calcium in cultured vascular smooth muscle cells of spontaneously hypertensive rats and Wistar-Kyoto rats / 中国结合医学杂志

<p><b>OBJECTIVE</b>To explore the effects of total flavonoids of Hippophae rhamnoides L. (TFH), quercetin (Que) and isorhamnetin (Isor) on the intracellular free calcium ([Ca(2+)](i)) in vascular smooth muscle cells (VSMC) of spontaneously hypertensive rats (SHR) and Wistar-Kyoto rats (WKY).</p><p><b>METHODS</b>Fluo 3-acetoxymethylester (Fluo-3/AM) was used to observe the effects of TFH (100 mg/L) and its essential monomers, namely Que (10(-4) mol/L) and Isor (10(-4) mol/L) on changes of [Ca(2+)](i) in cultured SHR and WKY VSMC (abbr. to Ca-SHR & Ca-WKY) following exposure to high K(+), norepinephrine (NE) and angiotensin II (Ang II), and to compare with the effects of verapamil (Ver).</p><p><b>RESULTS</b>(1) TFH, Que and Isor had inhibitory effects on resting Ca-SHR (P < 0.05), but had no significant effects on Ca-WKY (P > 0.05). (2) High K(+) could increase Ca-SHR more significantly than Ca-WKY (P < 0.05); TFH, Que and Isor could inhibit the elevation of [Ca(2+)](i) induced by high K(+)-depolarization, with the effects similar to that of Ver, and the effect on Ca-SHR was more significant than that on Ca-WKY (P < 0.05). (3) NE and Ang II could increase Ca-SHR more significantly than Ca-WKY (P < 0.05), TFH, Que and Isor had remarkably inhibitory effect on the elevation of Ca-SHR and Ca-WKY induced by NE or Ang II. (4) In the absence of extracellular Ca(2+), TFH, Que and Isor also had certain inhibitory effect on Ca-SHR and Ca-WKY induced by NE, and the effect on the former was more significant than that on the latter (P < 0.05).</p><p><b>CONCLUSION</b>TFH, Que and Isor might decrease the levels of [Ca(2+)](i) in VSMCs by blocking both voltage-dependent calcium channels (VDC) and receptor-operated calcium channels (ROC) in physiological or pathological state, which may be one of the important mechanisms of their hypotensive and protective effects on target organs in patients with hypertension.</p>