Acteoside alleviates blood-brain barrier damage induced by ischemic stroke through inhibiting microglia HMGB1/TLR4/NLRP3 signaling.

Ischemic stroke (IS) can cause severe harm, inducing oxidative stress, inflammation, and pyroptotic death. IS treatment efficacy remains limited, and microglia are important regulators of IS-related blood-brain barrier (BBB) damage. It is thus vital that new therapeutic agents capable of targeting microglia be identified to treat IS-related damage to the BBB. Acteoside (ACT), which is a compound derived from Cistanche tubulosa (Schenk) Wight., offers promising bioactivity, but its ability to protect against central nervous system injury remains to be documented. To clarify the protective benefits and mechanisms through which ACT can protect against damage to the BBB, a rat middle cerebral artery occlusion (MCAO) model system was herein employed. These in vivo analyses demonstrated that ACT was able to significantly reduce cerebral infarct size while improving their neurological scores and altering neurotrophic and inflammatory factor release. RNA sequencing and molecular docking studies highlighted the ability of ACT to exert its protective benefits via the HMGB1/TLR4/NLRP3 axis. Western immunoblotting and immunofluorescent staining for tight junction proteins additionally confirmed the ability of ACT to preserve BBB integrity. The underlying mechanisms were then explored with an oxygen-glucose deprivation (OGD) model in vitro with BV2 cells. This strategy thus confirmed that the ability of ACT to suppress microglial inflammatory and pyroptotic activity was HMGB1/TLR4/NLRP3 pathway-dependent. These data thus offer novel evidence that ACT can protect against IS-related damage to the BBB through the abrogation of inflammatory and pyroptotic activity, underscoring its promise as a novel lead compound for the therapeutic treatment of IS.

Astrocytes and microglia-targeted Danshensu liposomes enhance the therapeutic effects on cerebral ischemia-reperfusion injury.

Cerebral ischemia-reperfusion injury (CI/RI) is the main cause of disability and death in stroke without satisfactory therapeutic effect. Inflammation mediated by activation of astrocytes and microglia is the main pathological mechanism of CI/RI. Danshensu (DSS) has been shown to exert anti-inflammatory effects against brain injury. However, limited by its poor cellular permeability and low bioavailability, it is still needed the new DSS preparations with the ability to cross the blood-brain barrier (BBB) and target inflammatory glial cells. In this study, we developed phosphatidylserine (PS) and transferrin (TF) modified liposomes carrying DSS (TF/PS/DSS-LPs) to improve the therapeutic efficacy against ischemic stroke. First, TF molecules targeted transferrin receptor (TfR) that is overexpressed in the BBB. Following the liposomes enter the brain, PS modification allowed the liposomes to target and bind to the overexpressed phosphatidylserine-specific receptors (PSRs) on the surface of astrocytes and microglia. Furthermore, it enhanced the uptake of TF/PS/DSS-LPs by astrocytes and microglia, while polarizing astrocytes from A1 to A2 and microglia from M1 to M2, reducing neuronal inflammation, and ultimately ameliorating cerebral ischemic injury. Thus, TF/PS/DSS-LPs could potentially serve as a promising strategy for the CI/RI treatment.

Unzipped chromosome-level genomes reveal allopolyploid nematode origin pattern as unreduced gamete hybridization.

The formation and consequences of polyploidization in animals with clonal reproduction remain largely unknown. Clade I root-knot nematodes (RKNs), characterized by parthenogenesis and allopolyploidy, show a widespread geographical distribution and extensive agricultural destruction. Here, we generated 4 unzipped polyploid RKN genomes and identified a putative novel alternative telomeric element. Then we reconstructed 4 chromosome-level assemblies and resolved their genome structures as AAB for triploid and AABB for tetraploid. The phylogeny of subgenomes revealed polyploid RKN origin patterns as hybridization between haploid and unreduced gametes. We also observed extensive chromosomal fusions and homologous gene expression decrease after polyploidization, which might offset the disadvantages of clonal reproduction and increase fitness in polyploid RKNs. Our results reveal a rare pathway of polyploidization in parthenogenic polyploid animals and provide a large number of high-precision genetic resources that could be used for RKN prevention and control.

Development and future prospects of the artificial urinary sphincter.

BACKGROUND: Urinary incontinence is a urinary disorder in which urine leaks out involuntarily. This disorder seriously affects the quality of life of patients. For patients with mild incontinence, conservative treatment and medication may be the ideal treatment modality, but for patients with severe incontinence, an artificial urinary sphincter is currently a better treatment option. METHODS: In order to design an ideal artificial urinary sphincter, this article first searched and collected literature based on Preferred Reporting Items for Systematic Reviews and Meta-Analyses searched strategy by selecting specific subject terms and reviewed the artificial urinary sphincters that are currently in the research stage based on different activation methods. RESULTS: In response to the deficiencies of the existing artificial urinary sphincter, this article discusses the future optimization of the artificial urinary sphincter from three aspects: individual improvement of the artificial urinary sphincter, engineering design elements, and optimization of the artificial urinary sphincter manufacturing process. CONCLUSIONS: The manufacture of an idealized artificial urinary sphincter capable of meeting clinical needs is of great importance to improve the quality of life of patients. However, this approach is a reasonable option to explore and should not be overestimated until further evidence is available.

Novel synergistic mechanism of 11-keto-ß-boswellic acid and Z-Guggulsterone on ischemic stroke revealed by single-cell transcriptomics.

Although strides have been made, the challenge of preventing and treating ischemic stroke continues to persist globally. For thousands of years, the natural substances Frankincense and Myrrh have been employed in Chinese and Indian medicine to address cerebrovascular diseases, with the key components of 11-keto-ß-boswellic acid (KBA) and Z-Guggulsterone (Z-GS) being the active agents. In this study, the synergistic effect and underlying mechanism of KBA and Z-GS on ischemic stroke were examined using single-cell transcriptomics. Fourteen cell types were identified in KBA-Z-GS-treated ischemic penumbra, and microglia and astrocytes account for the largest proportion. They were further re-clustered into six and seven subtypes, respectively. GSVA analysis reflected the distinct roles of each subtype. Pseudo-time trajectory indicated that Slc1a2 and Timp1 were core fate transition genes regulated by KBA-Z-GS. In addition, KBA-Z-GS synergistically regulated inflammatory reactions in microglia and cellular metabolism and ferroptosis in astrocytes. Most notably, we established an innovative drug-gene synergistic regulation pattern, and genes regulated by KBA-Z-GS were divided into four categories based on this pattern. Finally, Spp1 was demonstrated as the hub target of KBA-Z-GS. Taken together, this study reveals the synergistic mechanism of KBA and Z-GS on cerebral ischemia, and Spp1 may be the synergistic target for that. Precise drug development targeting Spp1 may offer a potential therapeutic approach for treating ischemic stroke.

Magnetically controlled artificial urinary sphincter: An overview from existing devices to future developments.

BACKGROUND: Urinary incontinence is a common clinical problem in the world today. Artificial urinary sphincter is a good treatment approach for severe urinary incontinence, which is designed to mimic the action of the human urinary sphincter and assist patients to regain urinary function. METHODS: There are many control methods based on artificial urinary sphincter, such as hydraulic control, electromechanical control, magnetic control, and shape memory alloy control. In this paper, the literature was first searched and documented based on PRISMA search strategy for selected specific subject terms. Then, a comparison of artificial urethral sphincters based on different control methods was conducted, and the research progress of magnetically controlled artificial urethral sphincters was reviewed, and their advantages and disadvantages were summarized. Finally, the design factors for the clinical application of magnetically controlled artificial urinary sphincter are discussed. RESULTS: As magnetic control allows for non-contact force transfer and does not generate heat, it is proposed that magnetic control may be one of the more promising control methods. The design of future magnetically controlled artificial urinary sphincters will need a variety of considerations, including the structural design of the device, manufacturing materials, manufacturing costs, and convenience. In addition, validation of the safety and effectiveness of the device and device management are equally important. CONCLUSIONS: The design of an ideal magnetically controlled artificial urinary sphincter is of great importance to enhance patient treatment outcomes. However, there are still great challenges to be faced for the clinical application of such devices.

Research on biomechanical compatibility for a novel artificial anal sphincter with constant force.

BACKGROUND: Existing artificial anal sphincter studies have shown that biomechanical compatibility problem between artificial anal sphincter and rectum caused by long-term morphological changes of the tissue surrounding the implanted prosthesis can lead to device failure or tissue ischemic necrosis. In this article, a mechanical artificial anal sphincter with constant force clamping is designed based on the superelasticity of shape memory alloys, which improved the biomechanical compatibility of implantable artificial anal sphincter. METHODS: Firstly, the anatomical structure and the biomechanical properties of the rectum are analyzed to obtain the size parameters and material parameters of the rectal model. Secondly, a novel artificial anal sphincter with constant force is designed to improve the biomechanical compatibility between the artificial sphincter and the rectum. Thirdly, the static analysis of artificial anal sphincter is carried out by finite element analysis. RESULTS: The simulation results show that the artificial anal sphincter can maintain a constant clamping force of 4 N within a certain variation range of intestinal tissue thickness, which verifies the constant force characteristic of the artificial anal sphincter. The constant clamping force of the artificial anal sphincter to the rectum is 4 N that is greater than the clamping force 3.99 N required to close the rectum, which verifies the effectiveness the artificial anal sphincter. The surface contact stress and the minimum principal stress of the rectum in the clamping state are less than the pressure threshold, which verifies the safety of the artificial anal sphincter. CONCLUSIONS: The novel artificial anal sphincter has better biomechanical compatibility and improves the mechanical match between artificial sphincter and intestinal tissue. This study may provide more reasonable and effective simulation data for in vivo experiments of artificial anal sphincter in future, which may provide theoretical and technical support for further research about clinical application of artificial anal sphincter.

Cistanche tubulosa alleviates ischemic stroke-induced blood-brain barrier damage by modulating microglia-mediated neuroinflammation.

ETHNOPHARMACOLOGICAL RELEVANCE: Ischemic stroke (IS) has both high morbidity and mortality. Previous research conducted by our group demonstrated that the bioactive ingredients of the traditional medicinal and edible plant Cistanche tubulosa (Schenk) Wight (CT) have various pharmacological effects in treating nervous system diseases. However, the effect of CT on the blood-brain barrier (BBB) after IS are still unknown. AIM OF THE STUDY: This study aimed to identify CT's curative effect on IS and explore its underlying mechanism. MATERIALS AND METHODS: IS injury was established in a rat model of middle cerebral artery occlusion (MCAO). Gavage administration of CT at dosages of 50, 100, and 200 mg/kg/day was carried out for seven consecutive days. Network pharmacology was used for predicting the pathways and potential targets of CT against IS, and subsequent studies confirmed the relevant targets. RESULTS: According to the results, both neurological dysfunction and BBB disruption were exacerbated in the MCAO group. Moreover, CT improved BBB integrity and neurological function and protected against cerebral ischemia injury. Network pharmacology revealed that IS might involve neuroinflammation mediated by microglia. Extensive follow-up studies verified that MCAO caused IS by stimulating the production of inflammatory factors and microglial infiltration. CT was found to influence neuroinflammation via microglial M1-M2 polarization. CONCLUSION: These findings suggested that CT may regulate microglia-mediated neuroinflammation by reducing MCAO-induced IS. The results provide theoretical and experimental evidence for the efficacy of CT therapy and novel concepts for the prevention and treatment of cerebral ischemic injuries.

Microbiome-metabolomics analysis reveals the potential effect of verbascoside in alleviating cognitive impairment in db/db mice.

Cognitive impairment is the main central nervous system complication of diabetes, affecting the quality of life of patients. Herba Cistanche is a homologous plant widely used as a health food and therapeutic drug. Verbascoside, a signature component of Herba Cistanche, has anti-diabetic and neuroprotective effects. However, it is quickly metabolized by the gut microbiota, and the mechanism of its neuroprotection and improvement of learning and memory remains unclear. We investigated the effectiveness and potential mechanisms of verbascoside on cognitive dysfunction in db/db mice using a 16S rRNA microbiome and serum metabolomics approach. We found that 12-week treatment with verbascoside significantly inhibited insulin resistance, reduced blood glucose and lipids, and improved cognitive deficits. In addition, verbascoside increased the gut microbiota diversity, improved intestinal dysbiosis, attenuated intestinal barrier disruption, reduced the levels of inflammatory factors, regulated the expression of the metabolites associated with cognitive function, and enhanced the central insulin sensitivity and hippocampal synaptogenesis signaling. We revealed that verbascoside induced the enrichment of Alistipes, Roseburia, and Intestinimonas in the gut, suppressed the abundance of Escherichia-Shigella, increased the serum levels of gamma-aminobutyric acid, L-glutamic acid, and L-lysine, and decreased taurine expression. Finally, a strong association between gut microbes, serum metabolites, and cognitive performance affected by verbascoside was observed. Our research suggests that alterations in gut microbes/metabolites are involved in the development of diabetic cognitive dysfunction, which is alleviated by verbascoside in the db/db mice through restructuring the gut microbiota composition, ameliorating diabetic metabolic disorders, and attenuating pathological brain damage.

Frankincense-Myrrh treatment alleviates neuropathic pain via the inhibition of neuroglia activation mediated by the TLR4/MyD88 pathway and TRPV1 signaling.

BACKGROUND: Neuroglia are important modulators of neuronal functionality, and thus play an integral role in the pathogenesis and treatment of neuropathic pain (NP). According to traditional Chinese medicine, Frankincense-Myrrh is capable of "activating blood and dissipating blood stasis", and as such these two biological compounds are commonly used to treat NP, however, the mechanisms underlying the efficacy of such treatment are unclear. PURPOSE: This study aimed to further elucidate the protective effects associated with the Frankincense-Myrrh treatment of NP. METHODS: A chronic sciatic nerve compression injury (CCI) model of NP was established, after which animals were gavaged with Frankincense, Myrrh, Frankincense-Myrrh, or the positive control drug pregabalin for 14 days. Network pharmacology approaches were used to identify putative pathways and targets associated with the Frankincense-Myrrh-mediated treatment of NP, after which these targets were subjected to in-depth analyses. The impact of TLR4 blockade on NP pathogenesis was assessed by intrathecally administering a TLR4 antagonist (LRU) or the MyD88 homodimerization inhibitory peptide (MIP). RESULTS: Significant alleviation of thermal and mechanical hypersensitivity in response to Frankincense and Myrrh treatment was observed in NP model mice, while network pharmacology analyses suggested that the pathogenesis of NP may be related to TLR4/MyD88-mediated neuroinflammation. Consistently, Frankincense-Myrrh treatment was found to reduce TLR4, MyD88, and p-p65 expression in spinal dorsal horn neuroglia from treated animals, in addition to inhibiting neuronal TRPV1 and inflammatory factor expression. Intrathecal LRU and MIP delivery were sufficient to alleviate thermal and mechanical hyperalgesia in these CCI model mice, with concomitant reductions in neuronal TRPV1 expression and neuroglial activation in the spinal dorsal horn. CONCLUSION: These data suggest that Frankincense-Myrrh treatment was sufficient to alleviate NP in part via inhibiting TLR4/MyD88 pathway and TRPV1 signaling activity. Blocking TLR4 and MyD88 activation may thus hold value as a means of treating NP.

Effect and Mechanisms of Quercetin for Experimental Focal Cerebral Ischemia: A Systematic Review and Meta-Analysis.

Quercetin, a naturally occurring flavonoid, is mainly extracted from tea, onions, and apples. It has the underlying neuroprotective effect on experimental ischemic stroke. A systematic review and meta-analysis were used to assess quercetin's efficacy and possible mechanisms in treating focal cerebral ischemia. Compared with the control group, twelve studies reported a remarkable function of quercetin in improving the neurological function score (NFS) (P < 0.05), and twelve studies reported a significant effect on reducing infarct volume (P < 0.05). Moreover, two and three studies showed that quercetin could alleviate blood-brain barrier (BBB) permeability and brain water content, respectively. The mechanisms of quercetin against focal cerebral ischemia are diverse, involving antioxidation, antiapoptotic, anti-inflammation, and calcium overload reduction. On the whole, the present study suggested that quercetin can exert a protective effect on experimental ischemic stroke. Although the effect size may be overestimated because of the quality of studies and possible publication bias, these results indicated that quercetin might be a promising neuroprotective agent for human ischemic stroke. This study is registered with PROSPERO, number CRD 42021275656.

Component-target network and mechanism of Qufeng Zhitong capsule in the treatment of neuropathic pain.

ETHNOPHARMACOLOGICAL RELEVANCE: Qufeng Zhitong capsule (QFZTC) is a traditional Chinese medicine (TCM) clinically used for treating pain. However, the active ingredients of QFZTC and its pharmacological mechanism in the treatment of neuropathic pain (NP) remain unclear. AIM OF THE STUDY: We aimed to identify the active ingredients of QFZTC and reveal its target genes and underlying mechanism of action in NP. MATERIALS AND METHODS: High-performance liquid chromatography (HPLC) was used to identify the active ingredients of QFZTC. Network pharmacology analysis was conducted to determine the core targets and pathway enrichment of QFZTC. An NP mice model was established through chronic compression injury (CCI) surgery of the sciatic nerve, while von Frey instrumentation and a thermal stimulator were employed to measure the sensitivity of mice to mechanical and thermal stimuli. Immunofluorescence was used to observe the expression of TLR4 and p-P65 in microglia. Western blotting was used to detect the levels of protein expression of Iba-1, TLR4, MyD88, P65, p-P65, and c-Fos, while ELISA kits were used to detect the release of TNF-α, IL-6, and IL-1ß. RESULTS: Seven active ingredients were identified in QFZTC: gallic acid, loganylic acid, syringin, corilagin, loganin, ellagic acid, and osthole. Network analysis identified TLR4, TNF, IL6, IL1ß, and c-Fos as core targets, and Toll-like receptors and NF-κB as core signaling pathways. Treatment with QFZTC significantly relieved mechanical allodynia and thermal hyperalgesia in CCI mice models. CCI induced an increase in the expression of TLR4 and p-P65 in microglia, whereas QFZTC dose-dependently reduced the expression of Iba-1, TLR4, MyD88, and p-P65 in the spinal cord. QFZTC inhibited the expression of the c-Fos pain marker and reduced the expression of the TNF-α, IL-6, and IL-1ß inflammatory factors. CONCLUSION: We combined the active ingredients of QFZTC with network pharmacology research to clarify its biological mechanism in the treatment of NP. We demonstrated that QFZTC reduced NP in mice probably through regulating the spinal microglia via the TLR4/MyD88/NF-κB signaling pathway. Hence, QFZTC could be regarded as a potential drug for relieving NP.

LIFGO: A modular laser-induced fluorescence detection system based on plug-in blocks.

In this work, a laser-induced fluorescence (LIF) detection system built in a modular assembling mode was developed based on commercial LEGO blocks and 3D printed blocks. We designed and fabricated a variety of 3D printed building blocks fixed with optical components, including laser light source, filters, lens, dichroic mirror, photodiode detector, and control circuits. Utilizing the relatively high positioning precision of the plug-in blocks, a modular construction strategy was adopted using the flexible plug-in combination of the blocks to build a highly sensitive laser-induced fluorescence detection system, LIFGO. The LIFGO system has a simple structure which could be constructed by inexperienced users within 3 h. We optimized the structure and tested the performance of the LIFGO system, and its detection limits for sodium fluorescein solution in 100 µm i.d. and 250 µm i.d. capillaries were 7 nM and 0.9 nM, respectively. Based on the LIFGO system, we also built a simple capillary electrophoresis (CE) system and applied it to the analysis of DNA fragments to demonstrate its application possibility in biochemical analysis. The separation of 7 fragments in DL500 DNA markers were completed in 600 s. Because of the features of low cost (less than $100) and easy-to-build construction, we introduced the LIFGO system to the experimental teaching of instrumental analysis for undergraduate students. The modular construction form of the LIF detection system greatly reduces the threshold of instrument construction, which is conducive to the popularization of the LIF detection technique in routine laboratories as well as the reform of experimental teaching mode.

The decreased expression of integrin αv is involved in T-2 toxin-induced extracellular matrix degradation in chondrocytes.

T-2 toxin is one of the most toxic and common mycotoxins in grains and related products. It is considered a risk factor for Kashin-Beck disease (KBD), an endemic osteoarthritis. Both in vitro and in vivo studies have shown that T-2 toxin can cause extracellular matrix degradation; however, the underlying mechanism is unclear. Integrins have been found to regulate the expression of matrix metalloproteinases (MMPs), the 'scissors' of matrix proteins. In this study, we investigated whether integrin αv played a role in T-2 toxin-induced matrix degradation. Results from our study showed that the expression of integrin αv in the cartilage of rats fed T-2 toxin was reduced compared to that in rats fed a normal diet. Integrin αv was downregulated in T-2 toxin-treated C28/I2 chondrocytes, and selenium was found to have a protective effect. The expression of MMP-1, -3, -10, and -13 increased whereas that of type II collagen (Col II) protein decreased in C28/I2 cells treated with an integrin αv inhibitor. In conclusion, T-2 toxin can downregulate integrin αv expression in chondrocytes. Reduced integrin αv signalling could induce the release of MMPs, leading to matrix degradation.

Combination of Systems Pharmacology and Experimental Evaluation to Explore the Mechanism of Synergistic Action of Frankincense-Myrrh in the Treatment of Cerebrovascular Diseases.

Frankincense-Myrrh is a classic drug pair that promotes blood circulation, and eliminates blood stasis. The combination of the two drugs has a definite clinical effect on the treatment of cerebrovascular diseases (CBVDs), but its mechanism of action and compatibility have not been elucidated. In this study, the bioactive components, core targets, and possible synergistic mechanisms of Frankincense-Myrrh in the treatment of CBVDs are explored through systems pharmacology combined with in vivo and in vitro experiments. Comparing target genes of components in Frankincense and Myrrh with CBVD-related genes, common genes were identified; 15 core target genes of Frankincense-Myrrh for the treatment of CBVDs were then identified using protein-protein interaction (PPI) analysis. It was also predicted through Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis that the molecular mechanism of Frankincense-Myrrh action on CBVDs was mainly related to the regulation of neurotrophic factors and inflammatory responses. Frankincense-Myrrh significantly improved neurological function, decreased infarct volume, alleviated histopathological damage, inhibited microglial expression, and promoted the expression of neurons in middle cerebral artery occlusion (MCAO)-induced rats. The results of this study not only provide important theoretical support and experimental basis for the synergistic effect of Frankincense-Myrrh, but also provide new ideas for the prevention and treatment of cerebral ischemic injuries.

Decreased Expression of Heat Shock Protein 47 Is Associated with T-2 Toxin and Low Selenium-Induced Matrix Degradation in Cartilages of Kashin-Beck Disease.

Recent evidence suggests a role of type II collagen in Kashin-Beck disease (KBD) degeneration. We aimed to assess the abnormal expression of heat shock protein 47 (HSP47) which is associated with a decrease in type II collagen and an increase in cartilage degradation in KBD. Hand phalange cartilages were collected from KBD and healthy children. Rats were administered with T-2 toxin under the selenium (Se)-deficient diet. ATDC5 cells were seeded on bone matrix gelatin to construct engineered cartilaginous tissue. C28/I2 and ATDC5 cells and engineered tissue were exposed to different concentrations of T-2 toxin with or without Se. Cartilage degeneration was determined through histological evaluation. The distribution and expression of type II collagen and HSP47 were investigated through immunohistochemistry, western blotting, and real-time PCR. KBD cartilages showed increased chondronecrosis and extracellular matrix degradation in deep zone with decreased type II collagen and HSP47 expression. The low-Se + T-2 toxin animal group showed a significantly lower type II collagen expression along with decreased HSP47 expression. Decreased type II collagen and HSP47 in C28/I2 and ATDC5 cells induced by T-2 toxin showed a dose-dependent manner. Hyaline-like cartilage with zonal layers was developed in engineered cartilaginous tissues, with decreased type II collagen and HSP47 expression found in T-2 toxin-treated group. Se-supplementation partially antagonized the inhibitory effects of T-2 toxin in chondrocytes and cartilages. HSP47 plays a role in the degenerative changes of KBD and associated with T-2 toxin-induced decreased type II collagen expression, further promoting matrix degradation.

Reduced expression of α2 integrin is involved in T-2 toxin-induced matrix degradation in C28/I2 cells and cartilages from rats administrated with T-2 toxin.

T-2 toxin is a mycotoxin demonstrating several harmful effects on chondrocyte and cartilage functions. In the present study, we investigated the toxic effects of T-2 toxin on cartilage matrix degradation and evaluated the involvement of α2 integrin in T-2 toxin-induced matrix damage. In C28/I2 cells, T-2 toxin decreased cell viability in a dose-dependent manner. Regarding matrix degradation, T-2 toxin decreased type II collagen and increased matrix metalloproteinase 13 (MMP-13) expression. Moreover, T-2 toxin significantly decreased the expression of α2 integrin in C28/I2 cells, indicating impaired chondrocyte-matrix interaction. Additionally, cartilage matrix degradation with decreased type II collagen expression was observed in the animal model, established using rats treated with T-2 toxin, with or without a selenium-deficient diet, presenting chondrocytes with necrosis in the deep zone. Simultaneously, rats administered T-2 toxin demonstrated overtly decreased α2 integrin expression in the articular cartilage. In the T-2 toxin plus selenium-deficient diet group, α2 integrin expression was further decreased in the deep zone of the cartilage. Furthermore, inhibition of α2ß1 integrin in C28/I2 cells could induce MMP-13 activation and type II collagen reduction, contributing to matrix degradation. These results indicate that the cytotoxic effects of T-2 toxin on chondrocyte damage and cartilage matrix degradation are associated with α2 integrin downregulation, by reducing type II collagen and MMP-13 activation.

Integrin α2ß1 is involved in T-2 toxin-induced decrease of type II collagen in C28/I2 chondrocytes.

The T-2 toxin exerts a variety of toxic effects on both experimental animals and humans. The integrin family plays a major role in mediating cell-ECM interactions. Therefore, the present study aimed to investigate the involvement of integrin α2ß1 in T-2 toxin-induced C28/I2 chondrocyte damage. The pathological damage of articular cartilage injury induced by T-2 toxin was observed by H&E staining. The expression levels of collagen 2 and MMP-13 (Matrix metalloproteinases 13) were detected using immunohistochemistry in articular cartilage tissues and western blotting in the cells. The blocking effect of integrin α2ß1 inhibitor on T-2 toxin-induced chondrocyte matrix degradation was examined by western blotting. Further, the effect of integrin α2ß1 inhibitor on T-2 toxin-induced chondrocyte apoptosis was analyzed. About 100 ng/g body weight (BW)/day T-2 toxin was fed to Sprague-Dawley (SD) rats, T-2 toxin treatment (0, 6, 12, and 24 ng/mL) induced C28/I2 chondrocytes. Both in vivo and in vitro, chondrocyte survival was inhibited, and the production of type II collagen was significantly reduced (p < 0.05). However, the level of MMP-13 was up-regulated (p < 0.05). Matrix degradation was effectively blocked after the pre-treatment by integrin α2ß1 inhibitor (p < 0.05). Conclusively, Integrin α2ß1 is a critical signaling pathway for communication between cells and the extracellular matrix, the present study provides a new clue to elucidate the mechanism of T-2 toxin-induced chondrocyte damage.

JNK/AP-1 activation contributes to tetrandrine resistance in T-cell acute lymphoblastic leukaemia.

T-cell acute lymphoblastic leukaemia (T-ALL) is a challenging malignancy with a high relapse rate attributed to drug resistance. Tetrandrine (TET), a bisbenzylisoquinoline alkaloid extracted from a Chinese herb, is a potential anti-cancer and anti-leukaemic drug. In this study we investigated the mechanisms of TET resistance in T-ALL cells in vitro. Among the four T-ALL cell lines tested, Jurkat and CEM cells exhibited the lowest and highest resistance to TET with IC50 values at 24 h of 4.31±0.12 and 16.53±3.32 µmol/L, respectively. When treated with TET, the activity of transcription factor activator protein 1 (AP-1) was significantly decreased in Jurkat cells but nearly constant in CEM cells. To avoid cell-specific variation in drug resistance and transcription factor activities, we established a TET-R Jurkat subclone with the estimated IC50 value of 10.90±.92 µmol/L by exposing the cells to increasing concentrations of TET. Interestingly, when treated with TET, TET-R Jurkat cells exhibited enhanced AP-1 and NF-κB activity, along with upregulation of c-Jun N-terminal kinase (JNK) and extracellular signal-regulated kinase (ERK) signaling pathways, whereas the expression of P-gp was not altered. Selective inhibition of JNK but not ERK suppressed AP-1 activity and TET resistance in TET-R Jurkat cells and in CEM cells. These results demonstrate that Jurkat cells acquire TET resistance through activation of the JNK/AP-1 pathway but not through P-gp expression. The JNK/AP-1 pathway may be a potential therapeutic target in relapsed T-ALL.

Fluorescent coumarin-based probe for cysteine and homocysteine with live cell application.

Cysteine (Cys) and homocysteine (Hcy) are two of important biological thiols and function as important roles in several biological processes. The development of Cys and Hcy probes will help to explore the functions of biothiols in biological systems. In this work, a new coumarin-based probe AC, containing an acryloyl moiety, was developed for Cys and Hcy detection in cells. Cys and Hcy undergo a nucleophilic addition and subsequent cyclization reaction to remove to the acryloyl group and yield a fluorescent product, 7-hydroxylcomuarin. The probe AC showed good selectivity for cysteine and homocysteine over glutathione and other amino acids and had low detection limits of 65nM for Cys and 79nM for Hcy, respectively. Additionally, confocal imaging experiments demonstrated that the probe AC can be applied to visualize Cys and Hcy in living cells.