Caffeic acid alleviates cerebral ischemic injury in rats by resisting ferroptosis via Nrf2 signaling pathway.

There are few effective and safe neuroprotective agents for the treatment of ischemic stroke currently. Caffeic acid is a phenolic acid that widely exists in a number of plant species. Previous studies show that caffeic acid ameliorates brain injury in rats after cerebral ischemia/reperfusion. In this study we explored the protective mechanisms of caffeic acid against oxidative stress and ferroptosis in permanent cerebral ischemia. Ischemia stroke was induced on rats by permanent middle cerebral artery occlusion (pMCAO). Caffeic acid (0.4, 2, 10 mg·kg-1·d-1, i.g.) was administered to the rats for 3 consecutive days before or after the surgery. We showed that either pre-pMCAO or post-pMCAO administration of caffeic acid (2 mg·kg-1·d-1) effectively reduced the infarct volume and improved neurological outcome. The therapeutic time window could last to 2 h after pMCAO. We found that caffeic acid administration significantly reduced oxidative damage as well as neuroinflammation, and enhanced antioxidant capacity in pMCAO rat brain. We further demonstrated that caffeic acid down-regulated TFR1 and ACSL4, and up-regulated glutathione production through Nrf2 signaling pathway to resist ferroptosis in pMCAO rat brain and in oxygen glucose deprivation/reoxygenation (OGD/R)-treated SK-N-SH cells in vitro. Application of ML385, an Nrf2 inhibitor, blocked the neuroprotective effects of caffeic acid in both in vivo and in vitro models, evidenced by excessive accumulation of iron ions and inactivation of the ferroptosis defense system. In conclusion, caffeic acid inhibits oxidative stress-mediated neuronal death in pMCAO rat brain by regulating ferroptosis via Nrf2 signaling pathway. Caffeic acid might serve as a potential treatment to relieve brain injury after cerebral ischemia. Caffeic acid significantly attenuated cerebral ischemic injury and resisted ferroptosis both in vivo and in vitro. The regulation of Nrf2 by caffeic acid initiated the transcription of downstream target genes, which were shown to be anti-inflammatory, antioxidative and antiferroptotic. The effects of caffeic acid on neuroinflammation and ferroptosis in cerebral ischemia were explored in a primary microglia-neuron coculture system. Caffeic acid played a role in reducing neuroinflammation and resisting ferroptosis through the Nrf2 signaling pathway, which further suggested that caffeic acid might be a potential therapeutic method for alleviating brain injury after cerebral ischemia.

TIMP2 ameliorates blood-brain barrier disruption in traumatic brain injury by inhibiting Src-dependent VE-cadherin internalization.

Blood-brain barrier (BBB) disruption is a serious pathological consequence of traumatic brain injury (TBI), for which there are limited therapeutic strategies. Tissue inhibitor of metalloproteinase-2 (TIMP2), a molecule with dual functions of inhibiting MMP activity and displaying cytokine-like activity through receptor binding, has been reported to inhibit VEGF-induced vascular hyperpermeability. Here, we investigate the ability of TIMP2 to ameliorate BBB disruption in TBI and the underlying molecular mechanisms. Both TIMP2 and AlaTIMP2, a TIMP2 mutant without MMP-inhibiting activity, attenuated neurological deficits and BBB leakage in TBI mice; they also inhibited junctional protein degradation and translocation to reduce paracellular permeability in human brain microvascular endothelial cells (ECs) exposed to hypoxic plus inflammatory insult. Mechanistic studies revealed that TIMP2 interacted with α3ß1 integrin on ECs, inhibiting Src activation-dependent VE-cadherin phosphorylation, VE-cadherin/catenin complex destabilization, and subsequent VE-cadherin internalization. Notably, localization of VE-cadherin on the membrane was critical for TIMP2-mediated EC barrier integrity. Furthermore, TIMP2-mediated increased membrane localization of VE-cadherin enhanced the level of active Rac1, thereby inhibiting stress fiber formation. All together, our studies have identified an MMP-independent mechanism by which TIMP2 regulates EC barrier integrity after TBI. TIMP2 may be a therapeutic agent for TBI and other neurological disorders involving BBB breakdown.

Umbilical cord mesenchymal stem cell-conditioned medium inhibits microglial activation to ameliorate neuroinflammation in amyotrophic lateral sclerosis mice and cell models.

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease for which few effective therapeutic strategies are available. Increasing evidence indicates that neuroinflammation plays a significant role in ALS pathogenesis. Mesenchymal stem cell (MSC)-based therapy has been proposed for the treatment of neurodegenerative diseases, including ALS. In this study, we first demonstrated that systemic administration of conditioned medium derived from umbilical cord MSCs (UCMSC-CM) extends the lifespan of transgenic SOD1-G93A mice, a well-characterized model of familial ALS. Moreover, UCMSC-CM inhibits microglial activation and astrogliosis and alleviates the inflammatory milieu by reducing the release of proinflammatory cytokines and the expression of iNOS in the spinal cord. Using BV-2 cells overexpressing the SOD1-G93A mutant as an ALS cellular model, we uncovered that UCMSC-CM also suppresses the lipopolysaccharide (LPS)-induced inflammatory response, including reduced expression of proinflammatory cytokines and iNOS. Importantly, by culturing astrocytes alone in microglia-conditioned medium (MCM) or together with microglia in a transwell coculture system, we found that UCMSC-CM modulates the secretome of microglia exposed to inflammatory stimuli, thereby preventing the conversion of astrocytes to the A1 neurotoxic phenotype. This study revealed the anti-inflammatory properties of UCMSC-CM and its regulatory effect on glial activation in the treatment of neuroinflammation in ALS, providing strong evidence for the clinical application of UCMSC-CM.

Structural diversification of bioactive bibenzyls through modular co-culture leading to the discovery of a novel neuroprotective agent.

Bibenzyls, a kind of important plant polyphenols, have attracted growing attention for their broad and remarkable pharmacological activities. However, due to the low abundance in nature, uncontrollable and environmentally unfriendly chemical synthesis processes, these compounds are not readily accessible. Herein, one high-yield bibenzyl backbone-producing Escherichia coli strain was constructed by using a highly active and substrate-promiscuous bibenzyl synthase identified from Dendrobium officinale in combination with starter and extender biosynthetic enzymes. Three types of efficiently post-modifying modular strains were engineered by employing methyltransferases, prenyltransferase, and glycosyltransferase with high activity and substrate tolerance together with their corresponding donor biosynthetic modules. Structurally different bibenzyl derivatives were tandemly and/or divergently synthesized by co-culture engineering in various combination modes. Especially, a prenylated bibenzyl derivative (12) was found to be an antioxidant that exhibited potent neuroprotective activity in the cellular and rat models of ischemia stroke. RNA-seq, quantitative RT-PCR, and Western-blot analysis demonstrated that 12 could up-regulate the expression level of an apoptosis-inducing factor, mitochondria associated 3 (Aifm3), suggesting that Aifm3 might be a new target in ischemic stroke therapy. This study provides a flexible plug-and-play strategy for the easy-to-implement synthesis of structurally diverse bibenzyls through a modular co-culture engineering pipeline for drug discovery.

Treatment of Superficial Mycoses Using Photodynamic Therapy: A Systematic Review and Meta-Analysis.

Objective: The purpose of this study is to evaluate the effectiveness and safety of photodynamic therapy (PDT) in treating superficial fungal infections, and provide reference for clinical application. Methods: In accordance with Population, Intervention, Comparator, and Outcome (PICO), the research question and keywords were formulated. Records published in English by PubMed, Embase, Cochrane Library, and Web of Science as of November 14, 2022 were retrieved, including the keywords "mycoses," "tinea," "photochemotherapy," etc. Besides, meta-analysis performed by STATA and PROSPERO registration code was CRD42022363448. Results: One thousand four hundred eighty-four records were identified and 18 articles involving 343 patients with superficial fungal infections were enrolled. The overall mycological cure rate of PDT is 55% [95% confidence interval (CI): 0.46-0.65]. The fungal cure rate using methylene blue (MB) as photosensitizer (PS) is 67% (95% CI: 0.55-0.79); using 5-aminolevulinic acid is 34% (95% CI: 0.21-0.47); and using methyl aminolevulinate is 56% (95% CI: 0.33-0.78). The fungal cure rate of moderate-to-severe onychomycosis according to Onychomycosis Severity Index is 60% (95% CI: 0.47-0.73) and that of moderate onychomycosis is 66% (95% CI: 0.56-0.76). It was observed that the treatment parameters did not follow the same standard across studies. The majority of the included studies were moderate to low biased. Conclusions: PDT, particularly using MB as PS, has a certain mycological cure rate and safety at treating superficial mycoses. Due to the insufficient number of studies on PDT in the treatment of superficial fungal infections and the small sample size of some studies, more studies with standardized PDT parameters, large sample size, and long follow-up periods are needed to prove that PDT has the potential to become an alternative to traditional antifungal therapy or to find a better combination between them.

Protective effect of human umbilical cord mesenchymal stem cell derived conditioned medium in a mutant TDP-43 induced motoneuron-like cellular model of ALS.

Amyotrophic lateral sclerosis (ALS) is a multi-factor neurodegenerative disease, characterized by the loss of motor neurons. TAR DNA-binding protein 43 (TDP-43) mutation, accumulation and aggregation, as well as oxidative stress are recognized as major pathological denominators and biochemical markers for ALS. Recently, human umbilical cord mesenchymal stem cell-derived conditioned medium (UC-CM) has been introduced to treat ALS patients. However, there is no research for the protective effect of UC-CM on the TDP-43 model of ALS. In this study, we evaluated the potential neuroprotective effect of UC-CM on a cellular ALS model expressing TDP-43mutant M337V, as well as its underlying mechanism. We found that 24 h UC-CM treatment could protect M337V expressing motor neurons by increasing cell viability and reducing LDH leakage. Furthermore, the aggregation of M337V, generation of ROS, malondialdehyde (MDA), 4-hydroxynonenal (4-HNE), protein carbonyl and 8-OHdG were also reduced by UC-CM, indicating that UC-CM protected cells by reducing oxidative damage. Moreover, UC-CM significantly increased the expression of nuclear Nrf2 and its downstream enzyme HO1. The Nrf2 translocation inhibitor ML385 could inhibit the effect of UC-CM on the cell viability and aggregate of M337V. Our results suggest that UC-CM protect cells against M337V expression by its strong antioxidative effect via Nrf-2/HO-1 axis activation.

Electroacupuncture Reduces Cerebral Hemorrhage Injury in Rats by Improving Cerebral Iron Metabolism.

Objective: To study the effects of electroacupuncture at Baihui and Dazhui points on the expression of hepcidin (Hepc), transferrin (Tf), transferrin receptor (TfR), and ferritin (Ft) in rats with cerebral hemorrhage to provide a theoretical basis for the treatment of cerebral hemorrhage with acupuncture. Method: The model of cerebral hemorrhage in rats was established by autologous blood injection method and treated by electroacupuncture (EA) at the acupoints of Baihui and Dazhui. Hepc siRNA was injected into the lateral ventricle 30 min before model preparation to produce the cerebral hemorrhage model. The modified neurological severity score (mNSS) was used to assess the neurological function, and the total iron content in brain tissue was determined using atomic absorption spectrometry; the expression of Hepc, Ft, Tf, and TfR in perihematoma tissue was detected using immunohistochemistry; the interference efficiency of Hepc siRNA was detected using western blot and reverse transcription polymerase chain reaction (RT-PCR). Results: The degree of neurological deficit showed a downward trend at 3 days, 7 days, and 14 days, and electroacupuncture significantly reduced the neurological deficit score at each time point (P < 0.01). Regarding total iron content in brain tissue, on the 3rd day, the 7th day, and the 14th day, the iron content of the hematoma tissue after intracerebral hemorrhage was reduced by electroacupuncture (P < 0.01). Regarding immunohistochemical results. Hepc, Ft, Tf, and TfR protein expressions on day 14 were significantly higher after cerebral hemorrhage (P < 0.01). After electroacupuncture, the expression of Hepc, Ft, Tf, and TfR protein was significantly reduced (P < 0.01). Western blot and RT-PCR revealed that the interference efficiency of Hepc siRNA was statistically significant (P < 0.01). Conclusion: Electroacupuncture can reduce neurological severity scores in rats with cerebral hemorrhage and may exert cerebral protective effects by reducing Hepc protein and gene expression; lowering Ft, Tf, and TfR protein expression; and promoting iron metabolism in the brain of rats with cerebral hemorrhage.

Electroacupuncture Preconditioning Reduces Oxidative Stress in the Acute Phase of Cerebral Ischemia-Reperfusion in Rats by Regulating Iron Metabolism Pathways.

BACKGROUND: Oxidative stress is an important mechanism of cerebral ischemia-reperfusion injury. Ferroptosis caused by iron overload after cerebral ischemia-reperfusion is considered a common cause of oxidative stress. Many recent studies have shown that electroacupuncture (EA) can regulate the expression of inflammatory factors, and the use of electroacupuncture preconditioning can produce a protective effect, which can reduce injury after cerebral ischemia and reperfusion. We aimed to assess whether EA could be used to reduce oxidative stress. METHODS: The oxidative stress level of rats during the acute phase of cerebral ischemia and reperfusion was assessed with and without preconditioning with EA. Molecular biology methods were used to detect iron metabolism and oxidative stress-related proteins. RESULTS: Rats that had EA preconditioning had lower infarct volumes than rats in the control group. Furthermore, western blot analysis showed that the expression of iron metabolism-related protein FPN-1 was higher in the intervention group than in the model group after reperfusion. In this regard, further investigation also demonstrated higher expression of glutathione and glutathione peroxidase-4, and lower reactive oxygen species values in the brain tissue of the EA group were compared with those of the control group rats. CONCLUSIONS: Electroacupuncture preconditioning can reduce oxidative stress after cerebral ischemia-reperfusion by regulating iron overload.

Immunomodulatory effects of Hericium erinaceus derived polysaccharides are mediated by intestinal immunology.

This study was aimed at investigating the immunomodulating activity of Hericium erinaceus polysaccharide (HEP) in mice, by assessing splenic lymphocyte proliferation (cell-mediated immunity), serum hemolysin levels (humoral immunity), phagocytic capacity of peritoneal cavity phagocytes (macrophage phagocytosis), and NK cell activity. ELISA of immunoglobulin A (SIgA) in the lamina propria, and western blotting of small intestinal proteins were also performed to gain insight into the mechanism by which HEP affects the intestinal immune system. Here, we report that HEP improves immune function by functionally enhancing cell-mediated and humoral immunity, macrophage phagocytosis, and NK cell activity. In addition, HEP was found to upregulate the secretion of SIgA and activate the MAPK and AKT cellular signaling pathways in the intestine. In conclusion, all these results allow us to postulate that the immunomodulatory effects of HEP are most likely attributed to the effective regulation of intestinal mucosal immune activity.

Tunicamycin enhances human colon cancer cells to TRAIL-induced apoptosis by JNK-CHOP-mediated DR5 upregulation and the inhibition of the EGFR pathway.

Tumor necrosis factor related apoptosis-inducing ligand (TRAIL) is a cytokine that selectively induces apoptosis in many tumor cells while leaving normal cells intact and is thus an attractive candidate for antitumor therapies. This paper reports that the combination of tunicamycin plus TRAIL produced a strong synergistic effect in TRAIL-sensitive human colon cancer HCT116 cells and TRAIL-resistant HT-29 cells. On a cellular mechanistic level, tunicamycin-enhanced TRAIL-induced apoptosis by death receptor (DR) 5 upregulation and DR4 deglycosylation. Knockdown of DR5 but not DR4 expression by specific shRNAs or siRNAs significantly increased tunicamycin-mediated and TRAIL-mediated cell viability. DR5 induction was regulated by C/EBP homologous protein (CHOP) and JNK as CHOP siRNA or JNK inhibitor SP600125 considerably abolished the DR5 induction. In addition, tunicamycin inhibited epidermal growth factor receptor glycosylation and the downstream signaling pathways, Akt and extracellular signal-regulated kinases activation, which might also be required for TRAIL sensitization by tunicamycin. In summary, tunicamycin effectively enhanced TRAIL-induced apoptosis might through JNK-CHOP-mediated DR5 upregulation and the inhibition of the epidermal growth factor receptor pathway.