The Scutellaria-Coptis herb couple and its active small-molecule ingredient wogonoside alleviate cytokine storm by regulating the CD39/NLRP3/GSDMD signaling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: A drug pair is a fundamental aspect of traditional Chinese medicine prescriptions. Scutellaria baicalensis Georgi and Coptis chinensis Franch, commonly used as an herb couple (SBCC), are representative heat-clearing and dampness-drying drugs. They possess functions such as clearing heat, drying dampness, purging fire, and detoxifying. These herbs are used in both traditional and modern medicine for treating inflammation. AIM OF THE STUDY: This study investigated the effects of SBCC on cytokine storm syndrome (CSS) and explored its potential regulatory mechanism. MATERIALS AND METHODS: We assessed the impact of SBCC in a sepsis-induced acute lung injury mouse model by administering an intraperitoneal injection of LPS (15 mg/kg). The cytokine levels in the serum and lungs, the wet-to-dry ratio of the lungs, and lung histopathological changes were evaluated. The macrophages in the lung tissue were examined through transmission electron microscopy. Western blot was used to measure the levels of the CD39/NLRP3/GSDMD pathway-related proteins. Immunofluorescence imaging was used to assess the activation of pro-caspase-1 and ASC and their interaction. AMP-Glo™ assay was used to screen for active ingredients in SBCC targeting CD39. One of the ingredients was selected, and its effect on cell viability was assessed. We induced inflammation in macrophages using LPS + ATP and detected the levels of proinflammatory factors. The images of cell membrane large pores were captured using scanning electron microscopy, the interaction between NLRP3 and ASC was detected using immunofluorescence imaging, and the levels of CD39/NLRP3/GSDMD pathway-related proteins were assessed using Western blot. RESULTS: SBCC administration effectively mitigated LPS-induced cytokine storm, pulmonary edema and lung injury. Furthermore, it repressed the programmed death of lung tissue macrophages by inhibiting the NLRP3/GSDMD pyroptosis pathway and regulating the CD39 purinergic pathway. Based on the results of the AMP-Glo™ assay, we selected wogonoside for further valuation. Wogonoside alleviated LPS + ATP-induced inflammatory damage by regulating the inhibiting the NLRP3/GSDMD pyroptosis pathway and regulating the CD39 purinergic pathway. However, its effect on NLRP3 is not mediated though CD39. CONCLUSION: SBCC and its active small-molecule ingredient, wogonoside, improved CSS by regulating the NLRP3/GSDMD pyroptosis pathway and its upstream CD39 purinergic pathway. It is essential to note that the regulatory effect of wogonoside on NLRP3 is not mediated by CD39.

Genome-wide Analysis Identified SEMA4D, Novel Candidate Gene for Temperature Sensitivity in Patients With Non-Small Cell Lung Cancer.

BACKGROUND: In the era of precision medicine, individual temperature sensitivity has been highlighted. This trait has traditionally been used for cold-heat pattern identification to understand the inherent physical characteristics, which are influenced by genetic factors, of an individual. However, genome-wide association studies (GWASs) on this trait are limited. METHODS: Using genotype data from 90 patients with advanced non-small cell lung cancer (NSCLC) and epidermal growth factor receptor mutations, we performed a GWAS to assess the association between single nucleotide polymorphisms (SNPs) and temperature sensitivity, such as cold and heat scores. The score of each participant was evaluated using self-administered questionnaires on common symptoms and a 15-item symptom-based cold-heat pattern identification questionnaire. RESULTS: The GWAS was adjusted for confounding factors, including age and sex, and significant associations were identified for cold and heat scores: SNP rs145814326, located on the intron of SORCS2 at chromosome 4p16.1, had a P-value of 1.86 × 10-7; and SNP rs79297667, located upstream from SEMA4D at chromosome 9q22.2, had a P-value of 8.97 × 10-8. We also found that the genetic variant regulates the expression level of SEMA4D in the main tissues, including the lungs and white blood cells, in NSCLC. CONCLUSIONS: SEMA4D was found to be significantly associated with temperature sensitivity in patients with NSCLC, suggesting an increased expression of SEMA4D in patients with higher heat scores. The potential role of temperature sensitivity as a prognostic or predictive marker of immune response in NSCLC should be further studied.

Carnosol alleviates sepsis-induced pulmonary endothelial barrier dysfunction by targeting nuclear factor erythroid2-related factor 2/sirtuin-3 signaling pathway to attenuate oxidative damage.

Excessive reactive oxygen species production during acute lung injury (ALI) will aggravate the inflammatory process and endothelial barrier dysfunction. Carnosol is a natural phenolic diterpene with antioxidant and anti-inflammatory properties, but its role in treating sepsis-induced ALI remains unclear. This study aims to explore the protective effects and underlying mechanisms of carnosol in sepsis-induced ALI. C57BL/6 mouse were preconditioned with carnosol for 1 h, then the model of lipopolysaccharide (LPS)-induced sepsis was established. The degree of pulmonary edema, oxidative stress, and inflammation were detected. Endothelial barrier function was evaluated by apoptosis and cell junctions. In vitro, Mito Tracker Green probe, JC-1 staining, and MitoSOX staining were conducted to investigate the effect of carnosol on mitochondria. Finally, we investigated the role of nuclear factor-erythroid 2-related factor (Nrf2)/sirtuin-3 (SIRT3) in carnosol against ALI. Carnosol alleviated LPS-induced pulmonary oxidative stress and inflammation by inhibiting excess mitochondrial reactive oxygen species production and maintaining mitochondrial homeostasis. Furthermore, carnosol also attenuated LPS-induced endothelial cell barrier damage by reducing vascular endothelial cell apoptosis and restoring occludin, ZO-1, and vascular endothelial-Cadherin expression in vitro and in vivo. In addition, carnosol increased Nrf2 nuclear translocation to promote SIRT3 expression. The protective effects of carnosol on ALI were largely abolished by inhibition of Nrf2/SIRT3. Our study has provided the first evidence that the Nrf2/SIRT3 pathway is a protective target of the endothelial barrier in ALI, and carnosol can serve as a potential therapeutic candidate for ALI by utilizing its ability to target this pathway.

NTPDase1-ATP-P2Y2Rs axis in the sciatic nerve contributes to acupuncture at "Zusanli" (ST36)-induced analgesia in ankle arthritis rats.

BACKGROUND: The efficacy of acupuncture at Zusanli (ST36) in alleviating lower-limb pain is widely acknowledged in clinical practice, while its underlying mechanism remains incompletely elucidated. Our previous research had revealed that the prompt analgesia induced by needling-ST36 was accompanied by expression alterations in certain exco-nucleotidases within the sciatic nerve. Building upon this finding, the current work focused on NTPDase1, the primary ecto-nucleotidase in the human body, which converts ATP into AMP. METHODS: A 20-min acupuncture was administered unilaterally at the ST36 on rats with acute ankle arthritis. The pain thresholds of the injured hind paws were determined. Pharmacological interference was carried out by introducing the corresponding reagents to the sciatic nerve. ATP levels around the excised nerve were measured using a luciferase-luciferin assay. Live calcium imaging, utilizing the Fura 2-related-F340/F380 ratio, was conducted on Schwann cells in excised nerves and cultured rat SCs line, RSC96 cells. RESULTS: The analgesic effect induced by needling-ST36 was impaired when preventing ATP degradation via inhibiting NTPDase1 activities with ARL67156 or Ticlopidine. Conversely, increasing NTPDase1 activities with Apyrase duplicated the acupuncture effect. Similarly, preventing the conversion of AMP to adenosine via suppression of NT5E with AMP-CP hindered the acupuncture effect. Unexpectedly, impeded ATP hydrolysis ability and diminished NTPDase1 expression were observed in the treated group. Agonism at P2Y2Rs with ATP, UTP, or INS365 resulted in anti-nociception. Contrarily, antagonism at P2Y2Rs with Suramin or AR-C 118925xx prevented acupuncture analgesia. Immunofluorescent labeling demonstrated that the treated rats expressed more P2Y2Rs that were predominant in Schwann cells. Suppression of Schwann cells by inhibiting ErbB receptors also prevented acupuncture analgesia. Finally, living imaging on the excised nerves or RSC96 cells showed that agonism at P2Y2Rs indeed led to [Ca2+]i rise. CONCLUSION: These findings strongly suggest that the analgesic mechanism of needling-ST36 on the hypersensation in the lower limb partially relies on NTPDase1 activities in the sciatic nerve. In addition to facilitating adenosine signaling in conjunction with NT5E, most importantly, NTPDase1 may provide an appropriate low-level ATP milieu for the activation of P2Y2R in the sciatic nerve, particularly in Schwann cells.

Polymorphisms in the choline transporter SLC44A1 are associated with reduced cognitive performance in normotypic but not prenatal alcohol-exposed children.

BACKGROUND: Choline is essential for healthy cognitive development. Single nucleotide polymorphisms (SNPs; rs3199966(G), rs2771040(G)) within the choline transporter SLC44A1 increase risk for choline deficiency. In a choline intervention trial of children who experienced prenatal alcohol exposure (PAE), these alleles are associated with improved cognition. OBJECTIVE: This study aimed to determine if SNPs within SLC44A1 are differentially associated with cognition in children with PAE compared with normotypic controls (genotype × exposure). A secondary objective tested for an association of these SNPs and cognition in controls (genotype-only). DESIGN: This is a secondary analysis of data from the Collaborative Initiative on Fetal Alcohol Spectrum Disorders. Participants (163 normotypic controls, 162 PAE) underwent psychological assessments and were genotyped within SLC44A1. Choline status was not assessed. Association analysis between genotype × exposure was performed using an additive genetic model and linear regression to identify the allelic effect. The primary outcome was the interaction between SLC44A1 genotype × exposure status with respect to cognition. The secondary outcome was the cognitive-genotype association in normotypic controls. RESULTS: Genotype × exposure analysis identified 7 SNPs in SLC44A1, including rs3199966(G) and rs2771040(G), and in strong linkage (D' ≥ 0.87), that were associated (adjusted P ≤ 0.05) with reduced performance in measures of general cognition, nonverbal and quantitative reasoning, memory, and executive function (ß, 1.92-3.91). In controls, carriers of rs3199966(GT or GG) had worsened cognitive performance than rs3199966(TT) carriers (ß, 0.46-0.83; P < 0.0001), whereas cognitive performance did not differ by rs3199966 genotype in those with PAE. CONCLUSIONS: Two functional alleles that increase vulnerability to choline deficiency, rs3199966(G) (Ser644Ala) and rs2771040(G) (3' untranslated region), are associated with worsened cognition in otherwise normotypic children. These alleles were previously associated with greater cognitive improvement in children with PAE who received supplemental choline. The findings endorse that choline benefits cognitive development in normotypic children and those with PAE.

Histone deacetylase inhibitors inhibit lung adenocarcinoma metastasis via HDAC2/YY1 mediated downregulation of Cdh1.

Metastasis is a leading cause of mortality in patients with lung adenocarcinoma. Histone deacetylases have emerged as promising targets for anti-tumor drugs, with histone deacetylase inhibitors (HDACi) being an active area of research. However, the precise mechanisms by which HDACi inhibits lung cancer metastasis remain incompletely understood. In this study, we employed a range of techniques, including qPCR, immunoblotting, co-immunoprecipitation, chromatin-immunoprecipitation, and cell migration assays, in conjunction with online database analysis, to investigate the role of HDACi and HDAC2/YY1 in the process of lung adenocarcinoma migration. The present study has demonstrated that both trichostatin A (TSA) and sodium butyrate (NaBu) significantly inhibit the invasion and migration of lung cancer cells via Histone deacetylase 2 (HDAC2). Overexpression of HDAC2 promotes lung cancer cell migration, whereas shHDAC2 effectively inhibits it. Further investigation revealed that HDAC2 interacts with YY1 and deacetylates Lysine 27 and Lysine9 of Histone 3, thereby inhibiting Cdh1 transcriptional activity and promoting cell migration. These findings have shed light on a novel functional mechanism of HDAC2/YY1 in lung adenocarcinoma cell migration.

[Effect of catgut implantation on macrophage CD68, tumor necrosis factor-α and interleukin-1ß in "Zusanli" (ST 36) region of rats].

OBJECTIVE: To observe the expression of local macrophages and related cytokines tumor necrosis factor-α (TNF-α) and interleukin-1ß (IL-1ß) after catgut implantation in "Zusanli"(ST 36) in rats, so as to explore its underlying mechanisms in inducing therapeutic effect. METHODS: A total of 110 male SD rats were randomly divided into blank control group (n=10), catgut embedding (CE) group (n=50), and sham CE group (n=50). The CE and sham CE groups were randomly divided into 8 h, 3 d, 7 d, 14 d and 21 d subgroups after the intervention (n=10 in each time point group). Rats of the CE group were uniformly subjected into catgut embedding at ST36 once, and those of the sham CE group received embedding needle puncture at ST36 without catgut retention, and the blank control group was only grasped and fixed without other treatments. Tissues from the ST36 area in each group were collected at the corresponding time points, and the expression of CD68 in macrophages in the acupoint area was detected by immunofluorescence, the contents of TNF-α and IL-1ß in the acupoint area were detected by ELISA. RESULTS: Following catgut embedment at ST36, the contents of TNF-α and IL-1ß, and macrophage CD68 expression level began to increase at 8 h, peaked at 3 d, and then gradually decreased at 7, 14, and 21 d, being still higher in the CE group than in the blank control group at 21 d (P<0.05). Compared with the blank control group, the contents of TNF-α and IL-1ß, and macrophage CD68 expression were significantly increased at 8 h, and 3, 7, 14 and 21 d in the CE group (P<0.05). Following sham CE at ST36, the content of TNF-α at 8 h and 3 d, IL-1ß at 8 h and 3, 7 and 14 d, and expression of CD68 at 8 h were significantly increased in comparison with the blank control group (P<0.05). Comparison between the CE and sham CE groups showed that the contents of IL-1ß at 3, 7, 14 and 21 d, and contents of TNF-α，CD68 expression at 8 h, and 3, 7, 14 and 21 d were significantly higher in the CE group than in the sham CE group (P<0.05). CONCLUSION: Catgut embedding at ST36 can induce an increase levels of inflammatory cytokines TNF-α, IL-1ß and macrophage CD68 in the local microenvironment in rats, which may contribute to its functions in initiating therapeutic effect.

Curcumol Reduces Aerobic Glycolysis and Overcomes Chemoresistance by Inducing Cdh1-Mediated Skp2 Ubiquitination.

Colorectal cancer (CRC) is the third most common cancer worldwide. The main obstacle in treating advanced CRC is tumor recurrence and metastasis due to chemoresistance. S-phase kinase associated protein 2 (Skp2), an E3 ligase, is highly associated with tumor resistance and a poor prognosis. The results of immunoblotting, immunohistochemical staining, ubiquitination analysis, and co-immunoprecipitation (co-IP) assay revealed that the plant curcuma, curcumol, is a novel Skp2 inhibitor for CRC treatment. Curcumol inhibits aerobic glycolysis in CRC by inducing Skp2 degradation. Co-immunoprecipitation results showed that curcumol enhanced the interaction between cadherin-1 (Cdh1) and Skp2 and led to the ubiquitination and degradation of Skp2. Curcumol exhibited significant antitumor effects against CRC, such as increased intrinsic apoptosis and decreased tumorigenic properties, both in vivo and in vitro. Furthermore, curcumol overcame 5-fluorouracil (5-Fu) resistance in CRC and induced apoptosis in 5-Fu-resistant CRC cells. The present data revealed a novel antitumor mechanism of glycolytic regulation by curcumol, suggesting that curcumol may be a potential chemical candidate for treating 5-Fu-resistant CRC.

Micronutrient Supplementation and Bone Health After Prophylactic Total Gastrectomy in Patients With CDH1 Variants.

INTRODUCTION: Patients with germline variants in CDH1 who undergo prophylactic total gastrectomy (TG) are at risk of altered nutrient and drug absorption due to modified gastrointestinal anatomy. Bone mineral density loss and micronutrient deficiencies have not been described previously in this patient population. METHODS: In this study we included 94 patients with germline CDH1 variants who underwent prophylactic TG between October 2017 and February 2022. We examined pre- and post-gastrectomy bone mineral density (BMD); serum biomarkers including calcium, phosphorus, alkaline phosphatase, and 25 (OH)-vitamin D; and postoperative adherence to calcium and multivitamin supplementation. RESULTS: Almost all patients (92/94, 98%) lost a substantial amount of weight post-TG, with an average weight loss of 26.5% at 12 months post-surgery. Serum biomarkers of mineral metabolism, namely calcium and phosphorus, did not change significantly after TG. However, average BMD was decreased in all patients at 12 months post-TG. Nonadherence to calcium supplementation was associated with a decrease in BMD. Nonadherence to multivitamin supplementation was associated with greater percent BMD loss in the femoral neck and total hip. CONCLUSIONS: Appropriate micronutrient supplementation and nutritional counseling pre- and postoperatively in patients undergoing prophylactic TG are important to mitigate the long-term effects of gastrectomy on bone health.

The soluble CD83 protein prevents bone destruction by inhibiting the formation of osteoclasts and inducing resolution of inflammation in arthritis.

Here we show that soluble CD83 induces the resolution of inflammation in an antigen-induced arthritis (AIA) model. Joint swelling and the arthritis-related expression levels of IL-1ß, IL-6, RANKL, MMP9, and OC-Stamp were strongly reduced, while Foxp3 was induced. In addition, we observed a significant inhibition of TRAP+ osteoclast formation, correlating with the reduced arthritic disease score. In contrast, cell-specific deletion of CD83 in human and murine precursor cells resulted in an enhanced formation of mature osteoclasts. RNA sequencing analyses, comparing sCD83- with mock treated cells, revealed a strong downregulation of osteoclastogenic factors, such as Oc-Stamp, Mmp9 and Nfatc1, Ctsk, and Trap. Concomitantly, transcripts typical for pro-resolving macrophages, e.g., Mrc1/2, Marco, Klf4, and Mertk, were upregulated. Interestingly, members of the metallothionein (MT) family, which have been associated with a reduced arthritic disease severity, were also highly induced by sCD83 in samples derived from RA patients. Finally, we elucidated the sCD83-induced signaling cascade downstream to its binding to the Toll-like receptor 4/(TLR4/MD2) receptor complex using CRISPR/Cas9-induced knockdowns of TLR4/MyD88/TRIF and MTs, revealing that sCD83 acts via the TRIF-signaling cascade. In conclusion, sCD83 represents a promising therapeutic approach to induce the resolution of inflammation and to prevent bone erosion in autoimmune arthritis.

Cobalamin status is negatively correlated with vascular endothelial-cadherin in vegetarian and vegan women with vitamin B12 deficiency.

Optimal vitamin B12 status is important for vascular health. Vascular endothelial (VE) cadherin is an adherent junction protein involved in the maintenance of a functional endothelium. We hypothesized that vitamin B12 deficiency can negatively affect markers of vascular function, such as VE-cadherin. Within a human intervention study, we explored the possible association between cobalamin status (i.e., vitamin B12, holotranscobalamin, and homocysteine) and VE-cadherin (as marker of vascular health) in vegetarians/vegans (VEG) with B12 deficiency. The associations were evaluated at baseline and after 90-day supplementation with 2000 µg/wk of vitamin B12. On the whole, an inverse association between VE-cadherin and holotranscobalamin (P = .014) and a positive association between VE-cadherin and homocysteine (P = .041) was documented at baseline. VEG women showed higher levels of VE-cadherin compared with VEG men (P = .044), suggesting an increase in endothelial permeability. The intervention with vitamin B12 restored serum vitamin levels and improved the overall cobalamin status, whereas it did not affect VE-cadherin levels. The inverse association between holotranscobalamin and VE-cadherin was also maintained after intervention in women, corroborating the strong correlation between these 2 parameters. The results obtained seem to suggest a possible association between cobalamin status and VE-cadherin even if the intervention with B12 failed to positively affect VE-cadherin levels. Thus, further studies are needed to corroborate these findings and clarify the contribution of a vitamin B12 intervention on VE-cadherin levels in this target population. This trial was registered at ISRCTN registry (ISRCTN75099618).

Differential Expression of Platelet Activation Markers, CD62P and CD63, after Exposure to Breast Cancer Cells Treated with Kigelia Africana, Ximenia Caffra and Mimusops Zeyheri Seed Oils In Vitro.

Cancer patients, including breast cancer patients, live in a hypercoagulable state. Chemo- and hormone- therapy used in the treatment of breast cancer increases the risk of thrombosis. Due to differences in health care services between developed and developing countries, the survival rate of women with breast cancer in developing countries is low. Consequently, ethnomedicines are used and their efficacy as potential alternatives are being scientifically explored. The seed oils of Kigelia africana, Ximenia caffra and Mimusops zeyheri have anti-proliferative effects on hormone-dependent (MCF-7) and cytotoxic effects on hormone-independent (MDA-MB-231) breast cancer cells. In this study, we determined if these seed oils reduce the thrombogenic ability of breast cancer cells by measuring the platelet surface expression of the activation-specific antigens CD62P and CD63. MDA-MB-231 and MCF-7 cells were pretreated with the seed oils before being exposed to whole blood of human female volunteers. An increase in CD62P and CD63 expression following whole blood exposure to untreated breast cancer cells was observed. Treated MDA-MB-231 cells reduced CD62P and CD63 expression while treated MCF-7 cells increased CD62P and decreased CD63 expression. Kigelia africana, Ximenia caffra and Mimusops zeyheri seed oils are able to reduce the thrombogenic ability of MDA-MB-231 breast cancer cells.

Increased Ratio of CD14++CD80+ Cells/CD14++CD163+ Cells in the Infrapatellar Fat Pad of End-Stage Arthropathy Patients.

Objectives: This study sought to identify the ratio of M1/M2 cells in the infrapatellar fat pads (IFP) and subcutaneous fat tissues (SC) of osteoarthritis (OA) and rheumatoid arthritis (RA) patients. The clinical features of OA and RA patients treated with or without biological disease-modifying anti-rheumatic drugs (bDMARDs) were also assessed. Methods: IFP and SC were collected from patients with OA and RA who are undergoing total knee arthroplasty (TKA). CD14-positive cells were then isolated from these samples. Flow cytometry was used to determine the number of CD14++CD80+ cells and CD14++CD163+ cells. The expression levels of lipid transcription factors, such as sterol regulatory element-binding protein 1 (SREBP1) and liver X receptor alpha (LXRA), and inflammatory cytokines were also evaluated. Results: Twenty OA patients and 22 RA patients were enrolled in this study. Ten of the RA patients (45.4%) received bDAMRDs before TKA. On average, a fivefold increase in the number of CD14-positive cells and lower expression levels of SREBP1C and LXRA were observed in OA IFP relative to OA SC; however, these results were not obtained from the RA samples. The median ratio of CD14++CD80+ cells/CD14++CD163+ cells of OA IFP was 0.87 (0.76-1.09, interquartile range), which is higher to that of OA SC with a lower ratio (p = 0.05835). Conclusions: The quantity and quality of CD14-positive cells differed between IFP and SC in arthropathy patients. To our knowledge, this is the first study to characterize the ratio of M1/M2 cells in the IFP and SC of end-stage OA and RA patients. The increased ratio of CD14++CD80+ cells/CD14++CD163+ cells in the IFP from patients with OA and RA treated with bDMARDs indicated that inflammation was localized in the IFP. As adipose tissue-derived innate immune cells were revealed as one of the targets for regulating inflammation, further analysis of these cells in the IFP may reveal new therapeutic strategies for inflammatory joint diseases.

BST1 regulates nicotinamide riboside metabolism via its glycohydrolase and base-exchange activities.

Nicotinamide riboside (NR) is one of the orally bioavailable NAD+ precursors and has been demonstrated to exhibit beneficial effects against aging and aging-associated diseases. However, the metabolic pathway of NR in vivo is not yet fully understood. Here, we demonstrate that orally administered NR increases NAD+ level via two different pathways. In the early phase, NR was directly absorbed and contributed to NAD+ generation through the NR salvage pathway, while in the late phase, NR was hydrolyzed to nicotinamide (NAM) by bone marrow stromal cell antigen 1 (BST1), and was further metabolized by the gut microbiota to nicotinic acid, contributing to generate NAD+ through the Preiss-Handler pathway. Furthermore, we report BST1 has a base-exchange activity against both NR and nicotinic acid riboside (NAR) to generate NAR and NR, respectively, connecting amidated and deamidated pathways. Thus, we conclude that BST1 plays a dual role as glycohydrolase and base-exchange enzyme during oral NR supplementation.

Shikonin alleviates choroidal neovascularization by inhibiting proangiogenic factor production from infiltrating macrophages.

Choroidal neovascularization (CNV), a feature of neovasular age-related macular degeneration (AMD), acts as a leading cause of vision loss in the elderly. Shikonin (SHI), a natural bioactive compound extracted from Chinese herb radix arnebiae, exerts anti-inflammatory and anti-angiogenic roles and also acts as a potential pyruvate kinase M2 (PKM2) inhibitor in macrophages. The major immune cells macrophages infiltrate the CNV lesions, where the production of pro-angiognic cytokines from macrophage facilitates the development of CNV. PKM2 contributes to the neovascular diseases. In this study, we found that SHI oral gavage alleviated the leakage, area and volume of mouse laser-induced CNV lesion and inhibited macrophage infiltration without ocular cytotoxicity. Moreover, SHI inhibited the secretion of pro-angiogenic cytokine, including basic fibroblast growth factor (FGF2), insulin-like growth factor-1 (IGF1), chemokine (C-C motif) ligand 2 (CCL2), placental growth factor and vascular endothelial growth factor (VEGF), from primary human macrophages by down-regulating PKM2/STAT3/CD163 pathway, indicating a novel potential therapy strategy for CNV.

Evaluation of microglia in a rodent model of Parkinson's disease primed with L-DOPA after sub-anesthetic ketamine treatment.

Parkinson's disease (PD) is a neurodegenerative disease caused by the death of dopaminergic neurons in the substantia nigra pars compacta (SNpc), characterized by motor dysfunction. While PD symptoms are well treated with L-DOPA, continuous use can cause L-DOPA-induced dyskinesia (LID). We have previously demonstrated that sub-anesthetic ketamine attenuated LID development in rodents, measured by abnormal involuntary movements (AIMs), and reduced the density of maladaptive striatal dendritic mushroom spines. Microglia may play a role by phagocytosing maladaptive neuronal spines. In this exploratory study, we hypothesized that ketamine would prevent AIMs and change microglia ramified morphology - an indicator of a microglia response. Unilaterally 6-hydroxydopamine (6-OHDA)-lesioned rats were primed with daily injections of L-DOPA for 14 days, treated on days 0 and 7 for 10-hours with sub-anesthetic ketamine (i.p.), and we replicated that this attenuated LID development. We further extended our prior work by showing that while ketamine treatment did lead to an increase of striatal interleukin-6 in dyskinetic rats, indicating a modulation of an inflammatory response, it did not change microglia number or morphology in the dyskinetic striatum. Yet an increase of CD68 in the SNpc of 6-OHDA-lesioned hemispheres post-ketamine indicates increased microglia phagocytosis suggestive of a lingering microglial response to 6-OHDA injury in the SNpc pointing to possible anti-inflammatory action in the PD model in addition to anti-dyskinetic action. In conclusion, we provide further support for sub-anesthetic ketamine treatment of LID. The mechanisms of action for ketamine, specifically related to inflammation and microglia phagocytic functions, are emerging, and require further examination.

Phytochemicals in Cancer Immune Checkpoint Inhibitor Therapy.

The interaction of immune checkpoint molecules in the tumor microenvironment reduces the anti-tumor immune response by suppressing the recognition of T cells to tumor cells. Immune checkpoint inhibitor (ICI) therapy is emerging as a promising therapeutic option for cancer treatment. However, modulating the immune system with ICIs still faces obstacles with severe immunogenic side effects and a lack of response against many cancer types. Plant-derived natural compounds offer regulation on various signaling cascades and have been applied for the treatment of multiple diseases, including cancer. Accumulated evidence provides the possibility of efficacy of phytochemicals in combinational with other therapeutic agents of ICIs, effectively modulating immune checkpoint-related signaling molecules. Recently, several phytochemicals have been reported to show the modulatory effects of immune checkpoints in various cancers in in vivo or in vitro models. This review summarizes druggable immune checkpoints and their regulatory factors. In addition, phytochemicals that are capable of suppressing PD-1/PD-L1 binding, the best-studied target of ICI therapy, were comprehensively summarized and classified according to chemical structure subgroups. It may help extend further research on phytochemicals as candidates of combinational adjuvants. Future clinical trials may validate the synergetic effects of preclinically investigated phytochemicals with ICI therapy.

Systemic Administration of PTH Supports Vascularization in Segmental Bone Defects Filled with Ceramic-Based Bone Graft Substitute.

Non-unions continue to present a challenge to trauma surgeons, as current treatment options are limited, duration of treatment is long, and the outcome often unsatisfactory. Additionally, standard treatment with autologous bone grafts is associated with comorbidity at the donor site. Therefore, alternatives to autologous bone grafts and further therapeutic strategies to improve on the outcome and reduce cost for care providers are desirable. In this study in Sprague-Dawley rats we employed a recently established sequential defect model, which provides a platform to test new potential therapeutic strategies on non-unions while gaining mechanistic insight into their actions. The effects of a combinatorial treatment of a bone graft substitute (HACaS+G) implantation and systemic PTH administration was assessed by µ-CT, histological analysis, and bio-mechanical testing and compared to monotreatment and controls. Although neither PTH alone nor the combination of a bone graft substitute and PTH led to the formation of a stable union, our data demonstrate a clear osteoinductive and osteoconductive effect of the bone graft substitute. Additionally, PTH administration was shown to induce vascularization, both as a single adjuvant treatment and in combination with the bone graft substitute. Thus, systemic PTH administration is a potential synergistic co-treatment to bone graft substitutes.

Anti-Inflammatory Effect of Allicin Associated with Fibrosis in Pulmonary Arterial Hypertension.

Pulmonary arterial hypertension (PAH) is characterized by pulmonary vascular remodeling. Recent evidence supports that inflammation plays a key role in triggering and maintaining pulmonary vascular remodeling. Recent studies have shown that garlic extract has protective effects in PAH, but the precise role of allicin, a compound derived from garlic, is unknown. Thus, we used allicin to evaluate its effects on inflammation and fibrosis in PAH. Male Wistar rats were divided into three groups: control (CON), monocrotaline (60 mg/kg) (MCT), and MCT plus allicin (16 mg/kg/oral gavage) (MCT + A). Right ventricle (RV) hypertrophy and pulmonary arterial medial wall thickness were determined. IL-1ß, IL-6, TNF-α, NFκB p65, Iκß, TGF-ß, and α-SMA were determined by Western blot analysis. In addition, TNF-α and TGF-ß were determined by immunohistochemistry, and miR-21-5p and mRNA expressions of Cd68, Bmpr2, and Smad5 were determined by RT-qPCR. Results: Allicin prevented increases in vessel wall thickness due to TNF-α, IL-6, IL-1ß, and Cd68 in the lung. In addition, TGF-ß, α-SMA, and fibrosis were lower in the MCT + A group compared with the MCT group. In the RV, allicin prevented increases in TNF-α, IL-6, and TGF-ß. These observations suggest that, through the modulation of proinflammatory and profibrotic markers in the lung and heart, allicin delays the progression of PAH.

Single Dose of N-Acetylcysteine in Local Anesthesia Increases Expression of HIF1α, MAPK1, TGFß1 and Growth Factors in Rat Wound Healing.

In this study, we aimed to investigate the influence of N-acetylcysteine (NAC) on the gene expression profile, neoangiogenesis, neutrophils and macrophages in a rat model of incisional wounds. Before creating wounds on the backs of 24 Sprague-Dawley rats, intradermal injections were made. Lidocaine-epinephrin solutions were supplemented with 0.015%, 0.03% or 0.045% solutions of NAC, or nothing (control group). Scars were harvested on the 3rd, 7th, 14th and 60th day post-surgery. We performed immunohistochemical staining in order to visualize macrophages (anti-CD68), neutrophils (anti-MPO) and newly formed blood vessels (anti-CD31). Additionally, RT-qPCR was used to measure the relative expression of 88 genes involved in the wound healing process. On the 14th day, the number of cells stained with anti-CD68 and anti-CD31 antibodies was significantly larger in the tissues treated with 0.03% NAC compared with the control. Among the selected genes, 52 were upregulated and six were downregulated at different time points. Interestingly, NAC exerted a significant effect on the expression of 45 genes 60 days after its administration. In summation, a 0.03% NAC addition to the pre-incisional anesthetic solution improves neovasculature and increases the macrophages' concentration at the wound site on the 14th day, as well as altering the expression of numerous genes that are responsible for the regenerative processes.