Berberine alleviates diabetic retinopathy by regulating the Th17/Treg ratio.

BACKGROUND: Diabetic retinopathy (DR) stands as a prominent complication of diabetes. Berberine (BBR) has reported to be effective to ameliorate the retinal damage of DR. Studying the potential immunological mechanisms of BBR on the streptozotocin (STZ) induced DR mouse model will explain the therapeutic mechanisms of BBR and provide theoretical basis for the clinical application of this drug. METHODS: C57BL/6 J mice were induced into a diabetic state using a 50 mg/(kg·d) dose of STZ over a 5-day period. Subsequently, they were subjected to a high-fat diet (HFD) for one month. Following a 5-week treatment with 100 mg/(kg·d) BBR, the concentrations of inflammatory factors in the mice's peripheral blood were determined using an enzyme-linked immunosorbent assay (ELISA). Hematoxylin-eosin staining was employed to scrutinize pathological changes in the mice's retinas, while flow cytometry assessed the proportions of T-lymphocyte subsets and the activation status of dendritic cells (DCs) in the spleen and lymph nodes. CD4+T cells and DC2.4 cell lines were utilized to investigate the direct and indirect effects of BBR on T cells under high glucose conditions in vitro. RESULTS: Following 5 weeks of BBR treatment in the streptozotocin (STZ) mouse model of DR, we observed alleviation of retinal lesions and a down-regulation in the secretion of inflammatory cytokines, namely TNF-α, IL-1ß, and IL-6, in the serum of these mice. And in the spleen and lymph nodes of these mice, BBR inhibited the proportion of Th17 cells and promoted the proportion of Treg cells, thereby down-regulating the Th17/Treg ratio. Additionally, in vitro experiments, BBR directly inhibited the expression of the transcription factor RORγt and promoted the expression of the transcription factor Foxp3 in T cells, resulting in a down-regulation of the Th17/Treg ratio. Furthermore, BBR indirectly modulated the Th17/Treg ratio by suppressing the secretion of TNF-α, IL-1ß, and IL-6 by DCs and enhancing the secretion of indoleamine 2,3-dioxygenase (IDO) and transforming growth factor-beta (TGF-ß) by DCs. This dual action inhibited Th17 cell differentiation while promoting Treg cells. CONCLUSION: Our findings indicate that BBR regulate T cell subpopulation differentiation, reducing the Th17/Treg ratio by directly or indirectly pathway. This represents a potential therapeutic avenue of BBR for improving diabetic retinopathy.

Identification of the normative use of medical protective equipment by fusion of object detection and keypoints detection.

BACKGROUND AND OBJECTIVE: Irregular use of personal protective equipment (PPE) seriously affects the occupational health and safety of healthcare workers, especially in large public health emergencies such as COVID-19. The danger often occurs in some complex scenarios where the use of certain equipment does not comply with the spatial rules it specifies. The single object detection-based method is difficult to effectively verify whether the worker's use of PPE is normative. Also detecting the use of six classes of PPE brings a large computational load to the task. METHODS: In this paper, we proposed an identification approach that combined human keypoints detection with deep learning object detection to help facilitate the monitoring of healthcare workers' standard PPE use. We used YOLOv4 as the baseline model for PPE detection and MobileNetv3 as the backbone of the detector to reduce the computational effort. In addition, High-Resolution Net (HRNet) was the benchmark for keypoints detection, characterizing the coordinates of 25 key points in the human body. Generalized Intersection over Union (GIoU) was used to establish the association between PPEs and key points, and by calculating the matching score between a PPE and the corresponding body bounding boxes, the authenticity of the PPE specification could be effectively inferred. RESULTS: The proposed detector is able to identify whether a healthcare worker is normatively using multiple equipment with a higher precision (95.81 %), recall (96.38 %), and F1-score (96.09 %). Meanwhile, the number of parameters (2.87 M) and the size of the model (6.4 MB) are also more lightweight than other comparative detectors. CONCLUSIONS: Our approach is more reliable for reasoning about the normality of personal protection for healthcare workers in some complex scenarios than a single object detection-based approach. The developed identification framework provides a new automated monitoring solution for protection management in healthcare, and the modular design brings more flexible applications for different medical operation scenarios.

Salidroside regulates tumor microenvironment of non-small cell lung cancer via Hsp70/Stub1/Foxp3 pathway in Tregs.

BACKGROUND: The treatment of non-small cell lung cancer (NSCLC) is challenging due to immune tolerance and evasion. Salidroside (SAL) is an extract in traditional Chinese medicine and has a potential antitumor effect. However, the mechanism of SAL in regulating the immunological microenvironment of NSCLC is yet to be clarified. METHODS: The mouse model with Lewis lung cancer cell line (3LL) in C57BL/6 mice was established. And then, the percentage of tumor-infiltrating T cell subsets including Treg was detected in tumor-bearing mice with or without SAL treatment. In vitro, the effect of SAL on the expression of IL-10, Foxp3 and Stub1 and the function of Treg were detected by flow cytometry. Network pharmacology prediction and molecular docking software were used to predict the target of SAL and intermolecular interaction. Furthermore, the effect of SAL on the expression of Hsp70 and the co-localization of Stub1-Foxp3 in Treg was confirmed by flow cytometry and confocal laser microscopy. Finally, Hsp70 inhibitor was used to verify the above molecular expression. RESULTS: We discovered that SAL treatment inhibits the growth of tumor cells by decreasing the percentage of tumor-infiltrated CD4+Foxp3+T cells. SAL treatment downregulates the expression of Foxp3 in Tregs, but increases the expression of Stub1, an E3 ubiquitination ligase upstream of Foxp3, and the expression of Hsp70. Inhibiting the expression of Hsp70 reverses the inhibition of SAL on Foxp3 and disrupts the colocalization of Stub1 and Foxp3 in the nucleus of Tregs. CONCLUSIONS: SAL inhibits tumor growth by regulating the Hsp70/stub1/Foxp3 pathway in Treg to suppress the function of Treg. It is a new mechanism of SAL for antitumor therapy.

Comparison of the Characteristics of Cytokine Storm and Immune Response Induced by SARS-CoV, MERS-CoV, and SARS-CoV-2 Infections.

Cytokine storm (CS) is a significant cause of death in patients with severe coronavirus pneumonia. Excessive immune-inflammatory reaction, many inflammatory cell infiltration, and extreme increase of proinflammatory cytokines and chemokines lead to acute lung injury and acute respiratory distress syndrome (ARDS). This review compares the characters of cytokine storms and immune responses caused by three highly pathogenic and infectious coronaviruses (HCoVs), including severe acute respiratory syndrome coronavirus (SARS-CoV), Middle East respiratory syndrome-coronavirus (MERS-CoV), and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), and analyzes the possible mechanisms to guide clinical treatment in the future.

Salidroside regulates DC through TLR4 to increase the lethality of T cells to lung cancer 3LLcells / 中国肿瘤生物治疗杂志

@#Objective： ：To investigatetheeffectofsalidroside(SAL)onthephenotype of dendritic cells (DCs) and the antitumor ability of cytotoxic T lymphocytes (CTL). Methods： ：Lewis lung cancer cell line 3LL, wild type (WT) C57BL/6 mice and TLR4-/- C57BL/6 mice were chosen for this study. Mice bone marrow derived DC precursor cells were obtained to differentiate into immature DCs, which were harvested on the sixth day of culture. CD11c+ DCs were obtained by magnetic beads screening, and further divided into PBS group, SAL group and lipopolysaccharide (LPS) group.After being cultured for 48 h, the effects of SAL on surface molecules and phagocytosis of DCs as well as the efffect of TLR4 pathway on the killing effect of T cells were detected by Fow cytometry. Results： ： Compared with PBS group, expressions of DC surface molecules CD80, CD86 and MHC Ⅱ significantly increased (all P<0.05), phagocytosis significantly decreased (P<0.05), and TLR4 expression level significantly increased (P<0.01) in SAL group; Compared with WT group, after being treated with SAL or LPS, the expressions of DC surface molecules CD80, CD86 and MHC Ⅱ decreased significantly in TLR4-/- group (all P<0.05); ComparedwithPBSgroup,theactivatedCTLinSALgroupexhibited a significantly elevated killing effect against lung cancer 3LLcells (P<0.05). Conclusion：SAL can induce DC maturation by regulating TLR4, thus improving the killing ability of T cells.

[Salidroside induces anti-tumor effect in dendritic cells via ERK pathway].

Objective To investigate the role of mitogen activated protein kinase/extracellular signal-regulated kinase (MAPK/ERK) pathway in the salidroside (SAL)-regulated antitumor immunity of dendritic cells (DCs). Methods Lewis lung cancer-bearing mouse model was established and treated with normal saline (NS) (0.2 mL/d), SAL [500 mg/(kg.d)] and cyclophosphamide (CTX) [10 mg/(kg.d)] in corresponding groups. Then their survival and tumor quality were recorded. Meanwhile, the bone marrow-derived DCs from the Lewis lung cancer-bearing mice were isolated in vitro and cultured for 7 days before collection. After sorted with magnetic beads, cells were co-cultured with PBS, SAL (0.05 mg/mL), LPS (200 ng/mL) or SAL combined with ERK inhibitor U0126 separately for 48 hours, followed by detection of the expression of phosphorylated ERK (p-ERK) by flow cytometry. In addition, after the stimulation of DCs by SAL, the expression of p38 MAPK and phosphorylated c-jun N-terminal kinase (p-JNK) was detected by flow cytometry. Moreover, after sorted with magneticbeads, DCs were co-cultured with PBS, SAL (0.05 mg/mL), LPS (200 ng/mL) or SAL combined with U0126. Then, the supernatant was collected, and IL-12p70 secretion ability of DCs was detected by ELISA. Finally, sensitized bone marrow-derived DCs from Lewis lung cancer-bearing mice were stimulated with SAL and co-cultured with spleen T lymphocytes purified by nylon fiber column from C57BL/6 mice as effector cells. 3LL target cells were then added at the target ratio of 10:1, 25:1, 50:1, followed by the detection of cell viability of cytotoxic T lymphocytes (CTLs) using CCK-8 assay. Results SAL could significantly inhibit tumor growth and improve the survival rate. Compared with PBS group, the phosphorylation of ERK protein on DC was enhanced significantly, and then reduced significantly after U0126 treatment. The phosphorylation of p38MAPK and c-jun N-terminal kinase (JNK) protein was not statistically affected by SAL. The level of IL-12p70 significantly increased in SAL group, and became remarkably higher after U0126 treatment. Finally, the killing ability of CTL was significantly enhanced by SAL. Conclusion SAL can inhibit the tumor growth of Lewis lung cancer-bearing mice and activate the ERK signaling pathway, thereby promoting IL-12p70 secretion in DCs and enhancing the cytotoxicity of CTL.

[Ethanol extract of Rhodiola rosea L. regulates the number of tumor infiltrating T cells to enhance antitumor effect in Lewis lung cancer-bearing mice].

Objective To explore the anti-tumor immune function of the Tibetan medicine Rhodiola rosea L. (RRL). Methods Lewis lung cancer-bearing mice were randomly divided into normal saline group, 500 mg/kg RRL ethanol extract treatment group, and 10 mg/kg cyclophosphamide (CTX) treatment group. All the groups underwent the treatment for 10 days. The mouse survival rate and tumor inhibitory rate were calculated. Additionally, the numbers of CD4+T and CD8+ T cells of tumor infiltrating lymphocytes as well as the proportion of FOXP3+ regulatory T cells (Tregs) in the CD4+CD25+Tregs were detected by flow cytometry. Besides, the serum levels of interleukin 2 (IL-2) and γ-interferon (IFN-γ) in the tumor-bearing mice were examined through ELISA, and the spleen cytotoxic T lymphocytes (CTL) activity was detected by the lactic dehydrogenase (LDH) release assay. Results Lewis tumor-bearing mice treated with the ethanol extract of RRL showed remarkably enhanced survival rate and inhibited tumor growth. Furthermore, the number of tumor infiltrating CD4+T and CD8+ T cells increased, while the proportion of FOXP3+ Tregs in the CD4+CD25+Tregs showed a declined tendency. Meanwhile, the serum IFN-γ and IL-2 levels in Lewis tumor-bearing mice increased, and the killing capacity of spleen CTL was enhanced. Conclusion The ethanol extract of RRL has a positive role in enhancing the anti-tumor immunity by regulating the number and function of immunocytes.