The long ß2,3-sheets encoded by redundant sequences play an integral role in the channel function of P2X7 receptors.

P2X receptors are a class of nonselective cation channels widely distributed in the immune and nervous systems, and their dysfunction is a significant cause of tumors, inflammation, leukemia, and immune diseases. P2X7 is a unique member of the P2X receptor family with many properties that differ from other subtypes in terms of primary sequence, the architecture of N- and C-terminals, and channel function. Here, we suggest that the observed lengthened ß2- and ß3-sheets and their linker (loop ß2,3), encoded by redundant sequences, play an indispensable role in the activation of the P2X7 receptor. We show that deletion of this longer structural element leads to the loss of P2X7 function. Furthermore, by combining mutagenesis, chimera construction, surface expression, and protein stability analysis, we found that the deletion of the longer ß2,3-loop affects P2X7 surface expression but, more importantly, that this loop affects channel gating of P2X7. We propose that the longer ß2,3-sheets may have a negative regulatory effect on a loop on the head domain and on the structural element formed by E171 and its surrounding regions. Understanding the role of the unique structure of the P2X7 receptor in the gating process will aid in the development of selective drugs targeting this subtype.

A conserved residue in the P2X4 receptor has a nonconserved function in ATP recognition.

Highly conserved amino acids are generally anticipated to have similar functions across a protein superfamily, including that of the P2X ion channels, which are gated by extracellular ATP. However, whether and how these functions are conserved becomes less clear when neighboring amino acids are not conserved. Here, we investigate one such case, focused on the highly conserved residue from P2X4, E118 (rat P2X4 numbering, rP2X4), a P2X subtype associated with human neuropathic pain. When we compared the crystal structures of P2X4 with those of other P2X subtypes, including P2X3, P2X7, and AmP2X, we observed a slightly altered side-chain orientation of E118. We used protein chimeras, double-mutant cycle analysis, and molecular modeling to reveal that E118 forms specific contacts with amino acids in the "beak" region, which facilitates ATP binding to rP2X4. These contacts are not present in other subtypes because of sequence variance in the beak region, resulting in decoupling of this conserved residue from ATP recognition and/or channel gating of P2X receptors. Our study provides an example of a conserved residue with a specific role in functional proteins enabled by adjacent nonconserved residues. The unique role established by the E118-beak region contact provides a blueprint for the development of subtype-specific inhibitors of P2X4.

GSK1702934A and M085 directly activate TRPC6 via a mechanism of stimulating the extracellular cavity formed by the pore helix and transmembrane helix S6.

Transient receptor potential canonical (TRPC) channels, as important membrane proteins regulating intracellular calcium (Ca2+i) signaling, are involved in a variety of physiological and pathological processes. Activation and regulation of TRPC are more dependent on membrane or intracellular signals. However, how extracellular signals regulate TRPC6 function remains to be further investigated. Here, we suggest that two distinct small molecules, M085 and GSK1702934A, directly activate TRPC6, both through a mechanism of stimulation of extracellular sites formed by the pore helix (PH) and transmembrane (TM) helix S6. In silico docking scanning of TRPC6 identified three extracellular sites that can bind small molecules, of which only mutations on residues of PH and S6 helix significantly reduced the apparent affinity of M085 and GSK1702934A and attenuated the maximal response of TRPC6 to these two chemicals by altering channel gating of TRPC6. Combing metadynamics, molecular dynamics simulations, and mutagenesis, we revealed that W679, E671, E672, and K675 in the PH and N701 and Y704 in the S6 helix constitute an orthosteric site for the recognition of these two agonists. The importance of this site was further confirmed by covalent modification of amino acid residing at the interface of the PH and S6 helix. Given that three structurally distinct agonists M085, GSK1702934A, and AM-0883, act at this site, as well as the occupancy of lipid molecules at this position found in other TRP subfamilies, it is suggested that the cavity formed by the PH and S6 has an important role in the regulation of TRP channel function by extracellular signals.

Altered allostery of the left flipper domain underlies the weak ATP response of rat P2X5 receptors.

Although the extracellular ATP-gated cation channel purinergic receptor P2X5 is widely expressed in heart, skeletal muscle, and immune and nervous systems in mammals, little is known about its functions and channel-gating activities. This lack of knowledge is due to P2X5's weak ATP responses in several mammalian species, such as humans, rats, and mice. WT human P2X5 (hP2X5Δ328-349) does not respond to ATP, whereas a full-length variant, hP2X5 (hP2X5-FL), containing exon 10 encoding the second hP2X5 transmembrane domain (TM2), does. However, although rat P2X5 (rP2X5) has a full-length TM2, ATP induces only weak currents in rP2X5, which prompted us to investigate the mechanism underlying this small ATP response. Here, we show that single replacements of specific rP2X5 residues with the corresponding residues in hP2X5 (S191F or F195H) significantly enhance the current amplitude of rP2X5. Using a combination of engineered disulfide cross-linking, single-channel recording, and molecular modeling, we interrogated the effects of S191F and F195H substitutions on the allostery of the left flipper (LF) domain. On the basis of our findings, we propose that the bound ATP-induced distinct allostery of the LF domain with that of other functional subtypes has caused the weak ATP response of rP2X5 receptors. The findings of our study provide the prerequisite for future transgenic studies on the physiological and pathological functions of P2X5 receptors.

Novel Role of vBcl2 in the Virion Assembly of Kaposi's Sarcoma-Associated Herpesvirus.

The viral Bcl-2 homolog (vBcl2) of Kaposi's sarcoma-associated herpesvirus (KSHV) displays efficient antiapoptotic and antiautophagic activity through its central BH3 domain, which functions to prolong the life span of virus-infected cells and ultimately enhances virus replication and latency. Independent of its antiapoptotic and antiautophagic activity, vBcl2 also plays an essential role in KSHV lytic replication through its amino-terminal amino acids (aa) 11 to 20. Here, we report a novel molecular mechanism of vBcl2-mediated regulation of KSHV lytic replication. vBcl2 specifically bound the tegument protein open reading frame 55 (ORF55) through its amino-terminal aa 11 to 20, allowing their association with virions. Consequently, the vBcl2 peptide derived from vBcl2 aa 11 to 20 effectively disrupted the interaction between vBcl2 and ORF55, inhibiting the incorporation of the ORF55 tegument protein into virions. This study provides new insight into vBcl2's function in KSHV virion assembly that is separable from its inhibitory role in host apoptosis and autophagy.IMPORTANCE KSHV, an important human pathogen accounting for a large percentage of virally caused cancers worldwide, has evolved a variety of stratagems for evading host immune responses to establish lifelong persistent infection. Upon viral infection, infected cells can go through programmed cell death, including apoptosis and autophagy, which plays an effective role in antiviral responses. To counter the host response, KSHV vBcl2 efficiently blocks apoptosis and autophagy to persist for the life span of virus-infected cells. Besides its anti-programmed-cell-death activity, vBcl2 also interacts with the ORF55 tegument protein for virion assembly in infected cells. Interestingly, the vBcl2 peptide disrupts the vBcl2-ORF55 interaction and effectively inhibits KSHV virion assembly. This study indicates that KSHV vBcl2 harbors at least three genetically separable functions to modulate both host cell death signaling and virion production and that the vBcl2 peptide can be developed as an anti-KSHV therapeutic application.

Effects of products designed to modulate the gut microbiota on hyperlipidaemia.

PURPOSE: Fatalities due to heart and cerebrovascular diseases caused by uncontrolled hyperlipidaemia increase every year; on the other hand, lipid-lowering drugs are known to cause side effects. The gut microbiota has been thoroughly investigated by researchers and consumers, because they have unique functional properties and littler side effects. However, the effects of the gut microbiota remain controversial. We conducted a meta-analysis to assess the effects of products designed to modulate the gut microbiota on various hyperlipidaemias. METHODS: We systematically searched PubMed, Embase, Cochrane Library (Central), and Web of Science for randomized controlled trials (published before June 2017, and those only in English) to compare treatment (products designed to modulate the gut microbiota) versus placebo. Our main endpoints were total cholesterol (TC), triglyceride (TG), high-density lipoprotein cholesterol (HDL-C), and low-density lipoprotein cholesterol (LDL-C) in serum. We assessed pooled data using a fixed effects model. RESULTS: Of 1337 identified studies, 21 were eligible and included in our analysis (n = 1436 participants). The combined estimate of effect size for the impact of products designed to modulate the gut microbiota on serum TC (WMD - 11.07 mg/dL, 95% CI - 13.72 to - 8.43, p < 0.001), LDL-C (WMD - 10.96 mg/dL, 95% CI - 13.37 to - 8.56, p < 0.001), and HDL-C (WMD 0.72 mg/dL, 95% CI 0.06-1.38, p = 0.032) were statistically significant, while no significant effect was found on TG concentrations (WMD - 0.56 mg/dL, 95% CI - 5.59 to 4.47, p = 0.828). Subgroup analysis showed parallel trials, probiotics, and long-term intervention had better effects on lowering blood lipid levels. CONCLUSION: Products designed to modulate the gut microbiota results in changes of the plasma lipid concentrations and these changes may protect against cardiovascular disease.

Gastroprotective Effect of Alkaloids from Cortex Phellodendri on Gastric Ulcers in Rats through Neurohumoral Regulation.

The present study aimed to investigate the gastroprotective activity of the total alkaloids from the bark of Phellodendron amurense and identify their possible mechanism. Total alkaloids were obtained through an alcohol extraction method and were analyzed using LC-MS/MS. Chronic gastric ulcers were induced by acetic acid (0.14 mol/L) filter paper on the gastric serosa. The antiulcer effect of total alkaloids was evaluated using the ulcer area, the ulcer inhibition ratio, and epidermal growth factor. The gastroprotective mechanism of total alkaloids was revealed using the levels of serotonin and noradrenaline. The results showed that oral administration of total alkaloids (30 mg/kg/day) obviously decreased the ulcer area (7.67 ± 2.06 mm2; p < 0.01) compared with the model group (15.15 ± 2.34 mm2). The ulcer inhibition ratio of the total alkaloids group (50 %) was higher than the omeprazole-treated group (46 %), which showed that the antiulcer effect of the total alkaloids may be superior to omeprazole. Besides, the total alkaloids significantly increased the epidermal growth factor level and accelerated the healing of ulcers. Histological examination of gastric tissues also supported the same conclusion. In addition, the total alkaloids significantly elevated the levels of serotonin and noradrenaline (both p < 0.01 compared to the model group). Our data indicates that total alkaloids of Cortex Phellodendri exerts a beneficial gastroprotective effect and the involved mechanism is likely neurohumoral regulation. Thus, Cortex Phellodendri may develop into a promising clinical medicinal agent for improving the quality of ulcer healing.

Monocyclic monoterpenes as penetration enhancers of ligustrazine hydrochloride for dermal delivery.

The purpose of this study was to explore the enhancing effects and the mechanism of monocyclic monoterpene penetration enhancers (menthol and menthone) on the transdermal absorption of ligustrazine hydrochloride (LH). Franz-type diffusion cells were used to determine percutaneous parameters of LH in vitro and surface changes of porcine skin were studied by a scanning electron microscope (SEM). The effects of promoters on the biophysical natures of stratum corneum (SC) were researched by Fourier transform-infrared (FT-IR). Penetration parameters of menthol (p < 0.01) and menthone groups (p < 0.05) were greater than those of the control; morphological changes of skin monitored by SEM demonstrated that the menthone group had the most disruption and desquamation of SC flakes, which resulted from extracted lipids. FT-IR measurements showed menthone had the greatest changes in peak shift and peak area, which resulted from C-H stretching vibrations of SC lipids. The results suggest that the penetration mechanism might include disturbing and extracting SC lipids and the hydrogen bond connection.

The In vitro/vivo Evaluation of Prepared Gastric Floating Tablets of Berberine Hydrochloride.

Currently available antiulcer drugs suffered from serious side effects which limited their uses and prompted the need for a safe and efficient new antiulcer agent. The objective of this project work was to retain the drug in the stomach for better antiulcer activity and less side effects. Hence, the aim of our present work was to prepare a gastric floating tablet of Berberine hydrochloride (Ber) with suitable in vitro/vivo properties. In this study, different Ber gastric floating tablets were prepared by simple direct compression using various amounts of HPMCK15M and Carbopol 971PNF combined with other tablet excipients. The properties of the tablets including hardness, buoyancy, swelling ability, in vitro drug release, and in vivo pharmacokinetic study were evaluated. The obtained results disclosed that hardness, floating, swelling, and in vitro drug release of the Ber tablets depended mainly on the ratio of polymer combinations. Moreover, among six formulations, F3 exhibited desirable floating, swelling, and extended drug release. In addition, in vivo pharmacokinetic study suggested that prepared gastric floating tablets had significantly sustained-releasing effects compared with market tablets. Therefore, the developed gastric floating tablets of Ber could be an alternative dosage form for treatment of gastrointestinal disease.

A Stromal Cell-Derived Factor 1α Analogue Improves Endothelial Cell Function in Lipopolysaccharide-Induced Acute Respiratory Distress Syndrome.

Endothelial cell (EC) dysfunction is a critical mediator of the acute respiratory distress syndrome (ARDS). Recent studies have demonstrated that stromal cell-derived factor 1α (SDF-1α) promotes EC barrier integrity. Our previous studies used a SDF-1α analogue CTCE-0214 (CTCE) in experimental sepsis and demonstrated that it attenuated vascular leak and modulated microRNA (miR) levels. We examined the hypothesis that CTCE improves EC function in lipopolysaccharide (LPS)-induced ARDS through increasing miR-126 expression. Human microvascular endothelial cells (HMVECs) were treated with thrombin to disrupt the EC integrity followed by incubation with CTCE or SDF-1α. Barrier function was determined by trans-endothelial electrical resistance assay. CTCE-induced alterations in miRNA expression and signaling pathways involved in barrier function were determined. Thrombin-induced vascular leak was abrogated by both CTCE and SDF-1α. CTCE also prevented thrombin-induced decreases of vascular endothelial (VE)-cadherin cell surface expression and expansion of the intercellular space. CTCE increased miR-126 levels and induced activation of AKT/Rac 1 signaling. Cotreatment with a miR-126 inhibitor blocked the protective effects of CTCE on AKT activation and endothelial permeability. In subsequent in vivo studies, ARDS was induced by intratracheal instillation of LPS. Intravenous injection of CTCE diminished the injury severity as evidenced by significant reductions in protein, immune cells, inflammatory cytokines and chemokines in the bronchoalveolar lavage fluid, increased miR-126 expression and decreased pulmonary vascular leak and alveolar edema. Taken together, our data show that CTCE improves endothelial barrier integrity through increased expression of miR-126 and activation of Rac 1 signaling and represents an important potential therapeutic strategy in ARDS.

Plasma levels of microRNA are altered with the development of shock in human sepsis: an observational study.

BACKGROUND: Endothelial dysfunction plays a critical role in the development of sepsis-related organ failure; however, the mechanisms that govern its development are not fully understood. Endothelial progenitor cells (EPCs) reduce vascular leak and organ failure in experimental sepsis while modulating plasma expression of microRNA (miRNA). MicroRNAs are small, noncoding segments of RNA that regulate gene expression and are known to modulate endothelial cell function and inflammatory signaling pathways. We hypothesized that miRNA may play an etiologic role in the endothelial dysfunction of sepsis and that their extracellular expression levels would be altered in those with shock. METHODS: Thirteen miRNAs were identified by literature search and analysis of the contents of human EPC-derived exosomes using real-time PCR. Plasma samples were obtained from patients within 24 hours of their admission to ICUs with severe sepsis (n = 62) and from healthy controls (n = 32) and real-time PCR was used to measure the expression of the candidate miRNAs. The Wilcoxon rank sum test was used to compare expression levels of the 13 candidate miRNAs in septic patients with (n = 29) and without (n = 33) shock while logistic regression was used to determine the area under the curve for associations between miRNA expression and shock. Bioinformatic analyses using miRNA databases were performed to identify pathways and gene targets of differentially expressed miRNA with potential relevance to sepsis-related shock. RESULTS: MiRNA-34a expression was significantly increased in the group who developed shock (p = 0.03) while miR-15a and miR-27a expressions were significantly decreased in this group (p = 0.006 and 0.03, respectively). The combined expression of these three miRNAs predicted shock with an area under the curve of 0.78 (95 % CI 0.66-0.90). In silico analyses predict that these three miRNAs regulate genes involved in endothelial cell cycle, apoptosis, VEGF signaling, LPS-stimulated MAPK signaling, and nuclear factor kappa B signaling. CONCLUSIONS: The plasma levels of miRNA are altered in patients with severe sepsis complicated by shock and may offer prognostic value as well as insights into the mechanisms of endothelial dysfunction in sepsis.

Anti-hyperuricemic and nephroprotective effects of rhein in hyperuricemic mice.

Hyperuricemia has been considered to be a key risk factor for kidney disease. The formation of uric acid crystals in the kidney further stimulates an intensive inflammatory response. Rhein possesses various pharmacological activities, including anti-inflammatory, antioxidative, antitumor, purgative effects, and so on. To our knowledge, no previous work has been reported about the therapeutic effect of rhein on urate nephropathy. In this study, a model of hyperuricemia and nephropathy induced by adenine and ethambutol in mice was established. Meanwhile, the potential beneficial effects and mechanisms of rhein on hyperuricemia and nephropathy were also investigated. The results demonstrated that rhein significantly decreased the serum uric acid level by inhibiting the xanthine oxidase activity and increasing the excretion of urinary uric acid. In addition, rhein also markedly improved kidney damage related to hyperuricemia. Further investigation indicated that rhein improved the symptoms of nephropathy through decreasing the production of proinflammatory cytokines, including interleukin 1ß, prostaglandin E2, and tumor necrosis factor-α and inhibiting the expression of transforming growth factor-ß1. The present study suggests that rhein may have a considerable potential for development as an anti-hyperuricemic and nephroprotective agent for clinical application.

Antioxidative Activity of Flavonoids from Abrus cantoniensis against Ethanol-Induced Gastric Ulcer in Mice.

The present study investigated the flavonoids from Abrus cantoniensis against ethanol-induced gastric ulcers in mice. The flavonoids from A. cantoniensis were extracted with ethanol and purified by macroporous resin and polyamide. The 2,2-diphenyl-1-picrylhydrazyl assay was used to measure the antioxidative activities in vitro. The ethanol-induced ulcer mouse model was used to evaluate the gastroprotective activities of the flavonoids from A. cantoniensis. In addition, a method was established to ensure accuracy for animal ulcer evaluation. The flavonoids from A. cantoniensis showed a strong free radical scavenging capacity with an IC50 of 43.83 µg/mL in the 2,2-diphenyl-1-picrylhydrazyl assay. At doses between 28.16-112.67 mg/kg, the flavonoids conspicuously reduced the ulcer index in ethanol-induced mice (p<0.001). Significant differences were found in the levels of superoxide dismutase, catalase, glutathione, and myeloperoxidase in the stomach tissues between the flavonoids from the A. cantoniensis groups and the ethanol control group. The gastroprotective effect of the flavonoids from A. cantoniensis could be due to its antioxidative activity of the defensive mechanism. The data revealed that the flavonoids from A. cantoniensis could be a potential therapeutic agent for gastric ulcer prevention and treatment.

Combined treatment with a CXCL12 analogue and antibiotics improves survival and neutrophil recruitment and function in murine sepsis.

Previous studies demonstrated that the CXCL12 peptide analogue CTCE-0214 (CTCE) has beneficial effects in experimental sepsis induced by cecal ligation and puncture (CLP). We examined the hypothesis that CTCE recruits neutrophils (PMN) to the site of infection, enhances PMN function and improves survival of mice in CLP-induced sepsis with antibiotic treatment. Septic mice (n=15) were administered imipenem (25mg/kg) and CTCE (10 mg/kg) subcutaneously vs. vehicle control at designated intervals post-CLP. CTCE treatment increased PMN recruitment in CLP-induced sepsis as evidenced by increased PMN in blood by 2.4±0.6 fold at 18h, 2.9±0.6 fold at 24h, respectively and in peritoneal fluid by 2.0±0.2 fold at 24h vs. vehicle control. CTCE treatment reduced bacterial invasion in blood (CFU decreased 77±11%), peritoneal fluid (CFU decreased 78±9%) and lung (CFU decreased 79±8% vs. CLP vehicle). The improved PMN recruitment and bacterial clearance correlated with reduced mortality with CTCE treatment (20% vs. 67% vehicle controls). In vitro studies support the notion that CTCE augments PMN function by enhancing phagocytic activity (1.25±0.02 fold), increasing intracellular production of ROS (32±4%) and improving bacterial killing (CFU decreased 27±3%). These composite findings support the hypothesis that specific CXCL12 analogues with ancillary antibiotic treatment are beneficial in experimental sepsis, in part, by augmenting PMN recruitment and function. This article is protected by copyright. All rights reserved.

Protective effects of oxymatrine on experimental diabetic nephropathy.

Diabetic nephropathy, one of the most common and serious vascular complications of both type 1 and type 2 diabetes mellitus, has become a major contributor of end-stage renal failure. The aims of this study were to investigate the effects and possible underlying action mechanism(s) of oxymatrine on renal damage in diabetic rats. Diabetes was induced in male Sprague-Dawley rats by administering a high-fat diet and an intraperitoneal 30 mg/kg streptozotocin injection. The animals were treated orally with saline, metformin hydrochloride, and oxymatrine at 50, 100, and 150 mg/kg/day for 11 weeks. At the end of the treatment, renal tissue, blood, and urine samples were collected for histological and biochemical examination. The results revealed that oxymatrine significantly decreased blood glucose, urinary protein and albumin excretion, serum creatinine, and blood urea nitrogen in diabetic rats, and ameliorated diabetes-induced glomerular and tubular pathological changes. Furthermore, oxymatrine significantly prevented oxidative stress and reduced the contents of renal advanced glycation end products, transforming growth factor-ß1, connective tissue growth factor, and inflammatory cytokines in diabetic rats. All these results indicate that oxymatrine has protective effects on experimental diabetic nephropathy by multiple mechanisms.

Structural insights into the orthosteric inhibition of P2X receptors by non-ATP analog antagonists.

P2X receptors are extracellular ATP-gated ion channels that form homo- or heterotrimers and consist of seven subtypes. They are expressed in various tissues, including neuronal and nonneuronal cells, and play critical roles in physiological processes such as neurotransmission, inflammation, pain, and cancer. As a result, P2X receptors have attracted considerable interest as drug targets, and various competitive inhibitors have been developed. However, although several P2X receptor structures from different subtypes have been reported, the limited structural information of P2X receptors in complex with competitive antagonists hampers the understanding of orthosteric inhibition, hindering the further design and optimization of those antagonists for drug discovery. We determined the cryogenic electron microscopy (cryo-EM) structures of the mammalian P2X7 receptor in complex with two classical competitive antagonists of pyridoxal-5'-phosphate derivatives, pyridoxal-5'-phosphate-6-(2'-naphthylazo-6'-nitro-4',8'-disulfonate) (PPNDS) and pyridoxal phosphate-6-azophenyl-2',5'-disulfonic acid (PPADS), and performed structure-based mutational analysis by patch-clamp recording as well as molecular dynamics (MD) simulations. Our structures revealed the orthosteric site for PPADS/PPNDS, and structural comparison with the previously reported apo- and ATP-bound structures showed how PPADS/PPNDS binding inhibits the conformational changes associated with channel activation. In addition, structure-based mutational analysis identified key residues involved in the PPNDS sensitivity of P2X1 and P2X3, which are known to have higher affinity for PPADS/PPNDS than other P2X subtypes.

A shared mechanism for TNP-ATP recognition by members of the P2X receptor family.

P2X receptors (P2X1-7) are non-selective cation channels involved in many physiological activities such as synaptic transmission, immunological modulation, and cardiovascular function. These receptors share a conserved mechanism to sense extracellular ATP. TNP-ATP is an ATP derivative acting as a nonselective competitive P2X antagonist. Understanding how it occupies the orthosteric site in the absence of agonism may help reveal the key allostery during P2X gating. However, TNP-ATP/P2X complexes (TNP-ATP/human P2X3 (hP2X3) and TNP-ATP/chicken P2X7 (ckP2X7)) with distinct conformations and different mechanisms of action have been proposed. Whether these represent species and subtype variations or experimental differences remains unclear. Here, we show that a common mechanism of TNP-ATP recognition exists for the P2X family members by combining enhanced conformation sampling, engineered disulfide bond analysis, and covalent occupancy. In this model, the polar triphosphate moiety of TNP-ATP interacts with the orthosteric site, while its TNP-moiety is deeply embedded in the head and dorsal fin (DF) interface, creating a restrictive allostery in these two domains that results in a partly enlarged yet ion-impermeable pore. Similar results were obtained from multiple P2X subtypes of different species, including ckP2X7, hP2X3, rat P2X2 (rP2X2), and human P2X1 (hP2X1). Thus, TNP-ATP uses a common mechanism for P2X recognition and modulation by restricting the movements of the head and DF domains which are essential for P2X activation. This knowledge is applicable to the development of new P2X inhibitors.

Coating urchinlike gold nanoparticles with polypyrrole thin shells to produce photothermal agents with high stability and photothermal transduction efficiency.

Photothermal therapy using inorganic nanoparticles (NPs) is a promising technique for the selective treatment of tumor cells because of their capability to convert the absorbed radiation into heat energy. Although anisotropic gold (Au) NPs present an excellent photothermal effect, the poor structural stability during storage and/or upon laser irradiation still limits their practical application as efficient photothermal agents. With the aim of improving the stability, in this work we adopted biocompatible polypyrrole (PPy) as the shell material for coating urchinlike Au NPs. The experimental results indicate that a several nanometer PPy shell is enough to maintain the structural stability of NPs. In comparison to the bare NPs, PPy-coated NPs exhibit improved structural stability toward storage, heat, pH, and laser irradiation. In addition, the thin shell of PPy also enhances the photothermal transduction efficiency (η) of PPy-coated Au NPs, resulting from the absorption of PPy in the red and near-infrared (NIR) regions. For example, the PPy-coated Au NPs with an Au core diameter of 120 nm and a PPy shell of 6.0 nm exhibit an η of 24.0% at 808 nm, which is much higher than that of bare Au NPs (η = 11.0%). As a primary attempt at photothermal therapy, the PPy-coated Au NPs with a 6.0 nm PPy shell exhibit an 80% death rate of Hela cells under 808 nm NIR laser irradiation.

Chronic cough relief by allosteric modulation of P2X3 without taste disturbance.

P2X receptors are cation channels that sense extracellular ATP. Many therapeutic candidates targeting P2X receptors have begun clinical trials or acquired approval for the treatment of refractory chronic cough (RCC) and other disorders. However, the present negative allosteric modulation of P2X receptors is primarily limited to the central pocket or the site below the left flipper domain. Here, we uncover a mechanism of allosteric regulation of P2X3 in the inner pocket of the head domain (IP-HD), and show that the antitussive effects of quercetin and PSFL2915 (our nM-affinity P2X3 inhibitor optimized based on quercetin) on male mice and guinea pigs were achieved by preventing allosteric changes of IP-HD in P2X3. While being therapeutically comparable to the newly licensed P2X3 RCC drug gefapixant, quercetin and PSFL2915 do not have an adverse effect on taste as gefapixant does. Thus, allosteric modulation of P2X3 via IP-HD may be a druggable strategy to alleviate RCC.

Hesperidin prevents retinal and plasma abnormalities in streptozotocin-induced diabetic rats.

Diabetic retinopathy is a complex disease that potentially involves increased production of advanced glycosylation end products (AGEs) and elevated aldose reductase (AR) activity, which are related with oxidative stress and inflammation. The aim of this study was to investigate the effects of hesperidin on retinal and plasma abnormalities in streptozotocin-induced diabetic rats. Hesperidin (100, 200 mg/kg daily) was given to diabetic rats for 12 weeks. The blood-retina breakdown (BRB) was determined after 2 weeks of treatment followed by the measurement of related physiological parameters with ELISA kits and immunohistochemistry staining at the end of the study. Elevated AR activity and blood glucose, increased retinal levels of vascular endothelial growth factor (VEGF), ICAM-1, TNF-α, IL-1β and AGEs as well as reduced retina thickness were observed in diabetic rats. Hesperidin treatment significantly suppressed BRB breakdown and increased retina thickness, reduced blood glucose, AR activity and retinal TNF-α, ICAM-1, VEGF, IL-1β and AGEs levels. Furthermore, treatment with hesperidin significantly reduced plasma malondialdehyde (MDA) levels and increased SOD activity in diabetic rats. These data demonstrated that hesperidin attenuates retina and plasma abnormalities via anti-angiogenic, anti-inflammatory and antioxidative effects, as well as the inhibitory effect on polyol pathway and AGEs accumulation.