Cholesterol neutralized vemurafenib treatment by promoting melanoma stem-like cells via its metabolite 27-hydroxycholesterol.

Vemurafenib has been used as first-line therapy for unresectable or metastatic melanoma with BRAFV600E mutation. However, overall survival is still limited due to treatment resistance after about one year. Therefore, identifying new therapeutic targets for melanoma is crucial for improving clinical outcomes. In the present study, we found that lowering intracellular cholesterol by knocking down DHCR24, the limiting synthetase, impaired tumor cell proliferation and migration and abrogated the ability to xenotransplant tumors. More importantly, administration of DHCR24 or cholesterol mediated resistance to vemurafenib and promoted the growth of melanoma spheroids. Mechanistically, we identified that 27-hydroxycholesterol (27HC), a primary metabolite of cholesterol synthesized by the enzyme cytochrome P450 27A1 (CYP27A1), reproduces the phenotypes induced by DHCR24 or cholesterol administration and activates Rap1-PI3K/AKT signaling. Accordingly, CYP27A1 is highly expressed in melanoma patients and upregulated by DHCR24 induction. Dafadine-A, a CYP27A1 inhibitor, attenuates cholesterol-induced growth of melanoma spheroids and abrogates the resistance property of vemurafenib-resistant melanoma cells. Finally, we confirmed that the effects of cholesterol on melanoma resistance require its metabolite 27HC through CYP27A1 catalysis, and that 27HC further upregulates Rap1A/Rap1B expression and increases AKT phosphorylation. Thus, our results suggest that targeting 27HC may be a useful strategy to overcome treatment resistance in metastatic melanoma.

[Brain Fog: A Concept Analysis].

Brain fog is a symptom that has gained increasing attention worldwide since the start of the COVID-19 pandemic, as patients affected by COVID-19 may experience cognitive dysfunction, colloquially known as brain fog, for a period of time after recovery. Brain fog affects activities of daily living and work performance and has the potential to negatively impact society and the economy. However, a clear definition and concept analysis of brain fog is lacking in the literature. In this article, a concept analysis of brain fog is conducted using Walker and Avant's concept analysis steps to verify the source and definition of brain fog, clarify related concepts similar to brain fog, and establish the defining attributes, antecedents, and consequences of this condition. Model, borderline, contrary, and related cases are listed to illustrate and provide related empirical references in the literature. The authors hope this article will provide a clearer understanding of brain fog, which then may be applied in nursing clinical practice and future research to develop strategies and care methods for improving brain fog symptoms.

Establishment of an epithelioid sarcoma PDCs and PDX to evaluate drug sensitivity.

Epithelioid sarcoma (ES) is a very rare mesenchymal malignancy. Its oncogenesis remains unknown, and few therapies are available for advanced patients. Improved therapies, such as combined therapies, are urgently needed for the treatment of advanced ES. To identify precision drugs for advanced ES patients, the sensitivity of the drugs must be screened and evaluated before they are used for treatment. In the present study, tumour tissue from an ES patient was subjected to NGS sequencing, cell culture and the construction of a PDX model. Using the early generations of tumour-derived cells, we performed a screen and found that the combined drugs ADM and trametinib exhibited coefficiency in tumour inhibition. This conclusion was further confirmed in experiments with later generations of cells and PDX models. Therefore, we suggest that early generations of tumour-derived cells be used to test the sensitivity of ES tumours to candidate drugs. Moreover, we speculate that trametinib may be combined with chemotherapy in ES patients to prolong the duration of the chemotherapeutic response.

The prognostic value of S-100 protein and Ki-67 index in olfactory neuroblastoma.

OBJECTIVE: To evaluate the prognostic value of S-100 protein and Ki-67 labeling index in olfactory neuroblastomas. METHODS: A retrospective study was conducted on a cohort of 85 patients with olfactory neuroblastomas. The immunohistochemical expression of S-100 and Ki-67 was assessed, and the predictive value of S-100 and Ki-67 was further evaluated. The optimal cutoff value of Ki-67 labeling index was determined using time-dependent receiver operating characteristic curve analysis. Overall survival and progression-free survival were assessed using the Kaplan-Meier method. RESULTS: A cut-off Ki-67 labeling index value of 67.5% was determined for prognosis in patients with olfactory neuroblastomas. There was a significant correlation between Ki-67 expression and cervical lymph node metastasis (P = 0.049). Compared with S-100 (+), S-100 (-) was associated with a higher rate of lymph node metastasis and a higher level of Ki-67 (P = 0.007, < 0.001, respectively), as well as an advanced Kadish stage (P = 0.037). Survival analyses showed that patients with S-100 (+) had better 5-year overall survival than those with S-100 (-) (P = 0.028), and patients with both S-100 (+) and Ki-67 (<67.5%) had superior 5-year overall survival compared with all the other patients (P = 0.0225). CONCLUSION: Our findings suggest that S-100 combined with Ki-67 labeling index are reliable prognostic factors in patients with olfactory neuroblastomas.

Myosin1f-mediated neutrophil migration contributes to acute neuroinflammation and brain injury after stroke in mice.

BACKGROUND: During the acute stroke phase, neutrophils from the peripheral blood are first to arrive in the ischemic brain, which then attracts other immune cells that exacerbate neuroinflammation in the ischemic tissue. Myosin1f was reported to specifically mediate neutrophil migration in the peripheral tissues, but whether it plays a critical role in the neuroinflammatory response after ischemic stroke remains unknown. In this study, we aim to test the hypothesis that myosin1f-mediated neutrophil migration is critical in acute neuroinflammation induced by ischemic stroke. METHODS: Myosin1f -/- and wild type (WT) mice were subjected to transient middle cerebral artery occlusion (MCAO). To determine which cells determine myosin1f's transmigration ability, bone marrow transplantation, neutrophil depletion, and adoptive neutrophil transfer were performed. The myosin1f RNA level was assessed in peripheral neutrophils by reverse transcription polymerase chain reaction (RT-PCR) at 1 day and 3 days after stroke. The infiltrating neutrophils were quantified by immunofluorescence staining and FACS at 72 h after reperfusion. RESULTS: The myosin1f -/- mice had significantly smaller infarctions than the myosin1f +/+ mice. Bone marrow transplantation from myosin1f -/- mice to recipient mice also had smaller infarctions compared to animals receiving bone marrow from myosin1f +/+ mice. By performing neutrophil depletion and adoptive transfer, we confirmed that myosin1f acts mainly in circulating neutrophils. RT-PCR showed that myosin1f gene expression was increased in the circulating blood neutrophils at 3 days after ischemia. The confocal immunostaining and FACS results confirmed that fewer neutrophils infiltrated into the ischemic brain in myosin1f -/- mice compared to WT mice. CONCLUSIONS: Myosin1f determines neutrophil migration into the ischemic hemisphere, which directly affects stroke outcome.

Clinical analysis of 90 cases of malignant lacrimal sac tumor.

PURPOSE: To investigate the pathology, clinical manifestations, and potential risk factors associated with the prognosis of malignant lacrimal sac tumors. In addition, the treatment outcomes and complications were also evaluated. METHODS: Ninety cases of malignant lacrimal sac tumors were retrospectively analyzed at our hospital. Pathological classifications, clinical manifestations, risk factors, and follow-up time were documented. The outcomes and complications were evaluated and compared among the various treatment modalities. RESULTS: The median follow-up time was 50 months (range, 3-258 months). The 5-year overall survival (OS) and progression-free survival (PFS) for all cases were 85.7 and 77.9%, respectively. The 5-year OS and PFS for 69 cases of squamous cell carcinoma were 87.6 and 76.3%, and which were 80.4 and 72.4% for 21 cases of non-squamous cell carcinoma, respectively. There was no difference of 5-year OS and PFS between squamous cell carcinoma and non-squamous cell carcinoma (p = 0.350 and p = 0.946). Positive lymph node status was associated with worse OS (p < 0.001) and PFS (p = 0.020). For the 23.3% of cases (21/90) treated with the definitive radiotherapy, the outcomes were equivalent to that of surgery combined with radiotherapy, with the incidence of treatment-related visual acuity complication not being significant. The addition of chemotherapy to the treatment course had a marginal and non-significant improvement in OS and distant metastasis-free survival. CONCLUSIONS: Lymph node status was found to be a key factor for prognosis. Advanced tumors could benefit from multimodality treatment, with radiotherapy playing an important role. However, the role of chemotherapy requires further investigation.

Molecular identification of Neofabraea species associated with bull's-eye rot on apple using rolling-circle amplification of partial EF-1α sequence.

A rolling-circle amplification (RCA) method with padlock probes targeted on EF-1α regions was developed for rapid detection of apple bull's-eye rot pathogens, including Neofabraea malicorticis, N. perennans, N. kienholzii, and N. vagabunda (synonym: N. alba). Four padlock probes (PLP-Nm, PLP-Np, PLP-Nk, and PLP-Nv) were designed and tested against 28 samples, including 22 BER pathogen cultures, 4 closely related species, and 2 unrelated species that may cause serious apple decays. The assay successfully identified all the bull's-eye rot pathogenic fungi at the level of species, while no cross-reaction was observed in all target species and no false-positive reaction was observed with all strains used for reference. This study showed that the use of padlock probes and the combination of probe signal amplification by RCA provided an effective and sensitive method for the rapid identification of Neofabraea spp. The method could therefore be a useful tool for monitoring bull's-eye rot pathogens in port quarantine and orchard epidemiological studies.

Up-regulation of MIC19 promotes growth and metastasis of hepatocellular carcinoma by activating ROS/NF-κB signaling.

BACKGROUND: Mitochondrial malfunction has been well-recognized as a critical step in the pathogenesis of many types of diseases, including cancer. MIC19 is a core a subunit of the MICOS complex that plays a critical role in maintaining the normal function of mitochondria. However, the biological functions of MIC19 in human hepatocellular carcinoma (HCC) remain unclear. METHODS: The expression level of MIC19 in HCC was evaluated by bioinformatics analysis, quantitative real-time PCR and immunohistochemistry staining assays. Cell growth and metastasis experiments were used to assess the biological functions of MIC19 in HCC cells. FINDINGS: MIC19 expression was frequently upregulated in both human HCC specimens and cell lines, and its upregulation is closely associated with patients' survival. Results from loss-of-function and gain-of-function experiments demonstrated that knockdown of MIC19 significantly attenuated, while overexpression of MIC19 enhanced, the proliferation, colony formation, migration and invasion abilities of HCC cells. Mechanistically, we found that MIC19 has no effect on mitochondrial energy production, while activated ROS/NF-κB signaling, which was required for MIC19-promoted HCC growth and metastasis. INTERPRETATION: Our findings suggest that MIC19 play a critical oncogenic role in HCC, implying that MIC19 may serve as a potential therapeutic target in the treatment of HCC.

Establishment of Coloproctitis Cancer Model in Mice and Evaluation of Therapeutic Effect of Chinese Medicine.

Colorectal cancer (CRC) is a common malignancy of the digestive system and has become the third most common malignancy worldwide and the second leading cause of malignancy-related death. Ulcerative coloproctitis (UC) is a precancerous lesion, and UC-associated CRC (UC-CRC) is the most common subtype of CRC. Therefore, a reasonable UC-CRC model is the cornerstone and guarantee of new drug development. Traditional Chinese medicine (TCM) has been widely used in the treatment of UC-CRC due to its good efficacy. As a classic tonic prescription of TCM, Liujunzi decoction (LJZD) has been widely used in the treatment of UC-CRC. In this study, a UC-CRC model was established by combining azomethane and dextran sulfate sodium, and the LJZD was administered. The data confirmed that LJZD can effectively inhibit cancer transition in UC-CRC by using mouse body weight, colorectal length, pathological and inflammatory factors, colorectal barrier function, and cancer markers. This protocol provides a system for evaluating the efficacy of TCM in the prevention and treatment of UC-CRC.

Enhancing the Antitumor Immunity of T Cells by Engineering the Lipid-Regulatory Site of the TCR/CD3 Complex.

The engagement of the T-cell receptor (TCR) by a specific peptide-MHC ligand initiates transmembrane signaling to induce T-cell activation, a key step in most adaptive immune responses. Previous studies have indicated that TCR signaling is tightly regulated by cholesterol and its sulfate metabolite, cholesterol sulfate (CS), on the membrane. Here, we report a novel mechanism by which CS modulates TCR signaling through a conformational change of CD3 subunits. We found that the negatively charged CS interacted with the positively charged cytoplasmic domain of CD3ε (CD3εCD) to enhance its binding to the cell membrane and induce a stable secondary structure. This secondary structure suppressed the release of CD3εCD from the membrane in the presence of Ca2+, which in turn inhibited TCR phosphorylation and signaling. When a point mutation (I/A) was introduced to the intracellular immunoreceptor tyrosine-based activation motifs (YxxI-x6-8-YxxL) of CD3ε subunit, it reduced the stability of the secondary structure and regained sensitivity to Ca2+, which abolished CS-mediated inhibition and enhanced the signaling of the TCR complex. Notably, the I/A mutation could be applied to both murine and human TCR-T cell therapy to improve the antitumor efficacy. Our study reveals insights into the regulatory mechanism of TCR signaling and provides a strategy to functionally engineer the TCR/CD3 complex for T cell-based cancer immunotherapy.

Lactobacillus fermentum CECT5716 Alleviates the Inflammatory Response in Asthma by Regulating TLR2/TLR4 Expression.

Background: Asthma is a chronic disease, which is harmful to the health of the body and the quality of life. Supplementation of Lactobacillus can affect the immune environment of the lungs through the gut-lung axis. This study aimed to explore the potential regulatory targets of Lactobacillus to relieve inflammation in asthma and determine a new approach for improving asthma. Methods: A mouse ovalbumin (OVA)-induced model was constructed. OVA mice were supplemented with Lactobacillus fermentum CECT5716 by gavage. The gut microbiota composition of normal and OVA mice was analyzed using 16S ribosomal DNA identification. BALF, serum, lung tissues, and duodenal tissues were collected. Wright's staining was performed to determine the cell content of the alveolar lavage fluid. Hematoxylin-eosin staining, Masson staining, and periodic acid-Schiff staining were performed to observe the improvement in the lungs of OVA mice supplemented with Lactobacillus. Immunofluorescence was performed to measure the severity of the intestinal barrier leakage. Enzyme-linked immunosorbent assay was carried out to determine the expression levels of inflammatory cell factors, while quantitative reverse transcription-polymerase chain reaction and western blotting were performed to detect the levels of toll-like receptor 2 (TLR2)/TLR4 expression and cell adhesion factors. Results: Compared with Control mice, OVA mice exhibited malignant conditions, such as intestinal leakage and lung edema. After supplementation with Lactobacillus, the inflammatory cell content in the bronchoalveolar lavage fluid decreased, and the inflammatory response was alleviated. The level of TLR2/TLR4 expression was reduced. The inflammatory cell infiltration in the airway mucosa of OVA mice was improved, alveolar swelling was reduced and the basement membrane appeared thinner. Conclusion: The Lactobacillus inhibited the TLR2/TLR4 expression in OVA mice. Supplementation with Lactobacillus can alleviate the inflammatory response in OVA mice, inhibit pulmonary fibrosis, and treat asthma.

The role of 18F-FDG PET/CT in diagnosis and treatment evaluation for ocular adnexal mucosa-associated lymphoid tissue lymphoma.

OBJECTIVE: The purpose of this study is to evaluate the value of fluorine-18-fludeoxyglucose positron emission tomography (18F-FDG PET)/CT in the diagnosis and treatment evaluation of ocular adnexal mucosa-associated lymphoid tissue (MALT) lymphoma. METHODS: 70 patients with OAML who received radiotherapy were recruited in our study. All the patients had the 18F-FDG PET/CT examination before the treatment. We retrospectively reviewed the medical records, pathological reports, laboratory results, and imaging features of all patients. The associations between 18F-FDG PET/CT parameters and Epstein-Barr virus antibodies, treatment response, MRI data, and Ki-67 expression were investigated. RESULTS: The PET/CT scan indicated that 80% (56/70) of the patients showed orbital FDG avidity. The median level of maximum standardized uptake value (SUVmax) of the lesions was 4.65 ± 3.00 (range:1.2-13.5). 92.0% (46/50) of the mass-forming lesions showed 18F-FDG avidity, while only 50.0% (10/20) of the non-massive lesions had 18F-FDG avidity (χ2 = 13.23, p=0.01). The SUVmax in orbit, conjunctiva, and lacrimal gland lymphoma were 5.6, 2.9, and 3.7, respectively. A significant difference was identified of SUVmax among the three locations' lymphoma using one-way ANOVA analysis (F = 5.039, p = 0.01). After completion of radiotherapy, the complete remission rate was achieved in 30.8% (4/13) of the patients without 18F-FDG avidity, and 70.4% (38/54) in cases with 18F-FDG avidity (χ2 = 5.43, p = 0.02). The correlation between high Ki-67 score and 18F-FDG avidity was confirmed (χ2 = 3.916, p = 0.048); however, no significant correlation was found between the SUVmax and Ki-67 score of the lesions (p = 0.971). Three patients (3/70, 4.3%) were upregulated the stage via PET/CT. CONCLUSION: 18F-FDG PET/CT had some potential values in the diagnosis and assessment of treatment response in patients with OAML. ADVANCES IN KNOWLEDGE: The value of 18F-FDG PET/CT for patients with OAML.

Ontogeny of RORγt+ cells in the intestine of newborns and its role in the development of experimental necrotizing enterocolitis.

BACKGROUND: Neonates possess an immature and plastic immune system, which is a major cause of some diseases in newborns. Necrotizing enterocolitis (NEC) is a severe and devastating intestinal disease that typically affects premature infants. However, the development of intestinal immune cells in neonates and their roles in the pathological process of NEC have not been elucidated. RESULTS: We examined the ontogeny of intestinal lamina propria lymphocytes in the early life of mice and found a high percentage of RORγt+ cells (containing inflammatory Th17 and ILC3 populations) during the first week of life. Importantly, the proportion of RORγt+ cells of intestinal lamina propria further increased in both NEC mice and patients tissue than the control. Furthermore, the application of GSK805, a specific antagonist of RORγt, inhibited IL-17A release and ameliorated NEC severity. CONCLUSIONS: Our data reveal the high proportion of RORγt+ cells in newborn mice may directly contribute to the development of NEC.

A Single-Center Validation of the Accuracy of a Photoplethysmography-Based Smartwatch for Screening Obstructive Sleep Apnea.

BACKGROUND: Obstructive sleep apnea (OSA), the most common upper-airway disease, is closely associated with the risk of cardiovascular diseases. However, the early screening of OSA is a main challenge, relying on polysomnography (PSG) or home sleep apnea test (HSAT) in hospitals. Photoplethysmography (PPG) has been developed as a novel technology for screening of OSA, while the validation of PPG-based smart devices is limited compared to that for PSG or HSAT devices. OBJECTIVE: This study aimed to investigate the feasibility and validity of a PPG-based smartwatch in the screening of OSA. METHODS: A total of 119 patients were recruited from the Chinese People's Liberty Army General Hospital (Beijing, China). Among them, 20 patients were assessed for a whole-night sleep study by a smartwatch and PSG simultaneously, as well as 82 cases by a smartwatch and HSAT simultaneously. Using PSG or HSAT as the "gold standard", we compared the accuracy, sensitivity, specificity, negative predictive value (NPV), positive predictive value (PPV), and positive likelihood ratio (+LR) or negative likelihood ratio (-LR) at three apnea hypopnea index (AHI) levels: AHI≥5, AHI≥15, and AHI≥30. RESULTS: A total of 17/119 patients were excluded from the study due to the poor quality of PPG signals. Among the remaining cases, 83 patients were diagnosed with OSA. Compared to HSAT device, the accuracy, sensitivity, and specificity of the PPG-based smartwatch in predicting moderate-to-severe OSA patients (AHI≥15) were 87.9%, 89.7%, and 86.0%, respectively. Compared to PSG device, the accuracy, sensitivity, and specificity of the PPG-based smartwatch in predicting OSA in patients (AHI≥5) were 81.1%, 76.5%, and 100%, respectively. CONCLUSION: The PPG-based smartwatch outperformed in terms of detecting OSA; nevertheless, validation in a large-scale population is imperative. TRIAL REGISTRATION: Chinese Clinical Trial Registry of the International Clinical Trials Registry Platform of the World Health Organization ChiCTR-OOC-17014138; http://www.chictr.org.cn/showprojen.aspx?proj=24191.

Probiotics: their action against pathogens can be turned around.

Probiotics when applied in complex evolving (micro-)ecosystems, might be selectively beneficial or detrimental to pathogens when their prophylactic efficacies are prone to ambient interactions. Here, we document a counter-intuitive phenomenon that probiotic-treated zebrafish (Danio rerio) were respectively healthy at higher but succumbed at lower level of challenge with a pathogenic Vibrio isolate. This was confirmed by prominent dissimilarities in fish survival and histology. Based upon the profiling of the zebrafish microbiome, and the probiotic and the pathogen shared gene orthogroups (genetic niche overlaps in genomes), this consequently might have modified the probiotic metabolome as well as the virulence of the pathogen. Although it did not reshuffle the architecture of the commensal microbiome of the vertebrate host, it might have altered the probiotic-pathogen inter-genus and intra-species communications. Such in-depth analyses are needed to avoid counteractive phenomena of probiotics and to optimise their efficacies to magnify human and animal well-being. Moreover, such studies will be valuable to improve the relevant guidelines published by organisations such as FAO, OIE and WHO.

Molecular cloning and mRNA expression of peroxiredoxin gene in black tiger shrimp (Penaeus monodon).

The techniques of homology cloning and anchored PCR were used to clone the peroxiredoxin (Prx) gene from black tiger shrimp (Penaeus monodon). The full length cDNA of black tiger shrimp Prx (PmPrx) contained a 5' untranslated region (UTR) of 51 bp, an ORF (open reading frame) of 582 bp encoding a polypeptide of 193 amino acids with an estimated molecular mass of 22.15 kDa and a 3' UTR of 948 bp. Sequence comparison showed that PmPrx shared higher identities with Prx IVs than that with other isoforms of Prx, indicating PmPrx was a member of the Prx IV family. A quantitative reverse transcriptase Real-Time PCR (qRT-PCR) assay was developed to assess the mRNA expression of PmPrx in different tissues and the temporal expression of PmPrx in the hepatopancreas challenged by lipopolyssacharide (LPS). Higher-level mRNA expression of PmPrx was detected in the tissues of hepatopancreas, gonad and heart. The expression of PmPrx in the hepatopancreas was up regulated after stimulated by LPS. The results indicated that PmPrx was a constitutive and inducible expressed protein and could be induced by LPS.

[Evidenced-based practice: intraoperative prevention of sharps injuries].

Sharps injuries are a common occupational hazard in the operating room. This paper presents a conceptual map of sharps injuries prevention at the intraoperative level based on the evidence-based literature. The three outcome indicators identified include safety precaution, no sharps injuries, and competence. Five preventive interventions are suggested, including using hands-free techniques, wearing double-walled gloves, using blunt-tipped needles, providing education courses and making safety devices available. Recommendations in this paper may be used as a reference in developing a standard protocol for preventing sharps injuries in the operating room.

Tracheobronchopathia Osteochondroplastica: Five Cases Report and Literature Review.

Tracheobronchopathia osteochondroplastica (TO) is a rare disease. Here, we report 5 TO cases treated at our hospital. Bronchoscopy showed typical multiple firm and glossy nodules in all the 5 cases. Conservative treatment effectively alleviated the symptoms. Tracheobronchopathia osteochondroplastica is a manageable disease. Awareness in clinicians is critical to avoid unnecessary treatment in patients with TO.

Response Surface Optimization of Catechin Liposomes Preparation Using the Rapid Expansion of Supercritical Solution Process.

Phospholipid liposomes are a promising drug delivery system. Catechin, a hydrophilic drug, was used to prepare catechin liposomes through a modified rapid expansion of supercritical solution (RESS) process in this study. The influences of operation parameters (i.e., temperature, pressure, and mass ratio of liposomal materials to catechin) on the properties of the prepared liposomes were determined using the single-factor analysis. The process was further optimized by response surface methodology (RSM) based on the Box-Behnken design (BBD). The encapsulation efficiency (EE) values can be adequately predicted using the obtained equation. The maximum EE value can reach 61.36±0.68% under the optimal parameters (i.e., the expansion temperature, pressure, and p/c mass ratio were 56.34 °C, 19.99 MPa, and 5.99, respectively). The prepared liposomes can effectively protect and stabilize the loaded catechin effectively. In addition, the in vitro release study showed the slow and sustained release behavior of the catechin liposomes.