Dual drug nanoparticle synergistically induced apoptosis, suppressed inflammation, and protected autophagic response in rheumatoid arthritis fibroblast-like synoviocytes.

Rheumatoid arthritis (RA) is a chronic immune-mediated joint inflammatory disorder associated with aberrant activation of fibroblast-like synoviocytes (FLS). Recently, FLS gained importance due to its crucial role in RA pathogenesis, and thus, targeting FLS is suggested as an attractive treatment strategy for RA. FLS-targeted approaches may be combined with disease-modifying antirheumatic drugs (DMARDs) and natural phytochemicals to improve efficacy in RA control and negate immunosuppression. In this study, we assessed the therapeutic effectiveness of DD NP HG in primary RA-FLS cells isolated from the synovial tissue of FCA-induced RA rats. We observed that DD NP HG had good biosafety for healthy FLS cells and, at higher concentrations, a mild inhibitory effect on RA-FLS. The combination therapy (DD NP HG) of MTX NP and PEITC NE in RA-FLS showed a higher rate of apoptosis with significantly reduced LPS-induced expression of pro-inflammatory cytokines (TNF-α, IL-17A, and IL-6) in arthritic FLS. Further, the gene expression studies showed that DD NP HG significantly down-regulated the mRNA expression of IL-1ß, RANKL, NFATc1, DKK1, Bcl-xl, Mcl-1, Atg12, and ULK1, and up-regulated the mRNA expression of OPG, PUMA, NOXA and SQSTM1 in LPS-stimulated RA-FLS cells. Collectively, our results demonstrated that DD NP HG significantly inhibited the RA-FLS proliferation via inducing apoptosis, down-regulating pro-inflammatory cytokines, and further enhancing the expression of genes associated with bone destruction in RA pathogenesis. A nanotechnology approach is a promising strategy for the co-delivery of dual drugs to regulate the RA-FLS function and achieve synergistic treatment of RA.

Biocompatible quaternary pullulan functionalized 2D MoS2 glycosheet-based non-leaching and infection-resistant coatings for indwelling medical implants.

Medical implants are frequently used in medicine and reconstructive surgery to treat various pathological and anatomical conditions. However, over time, biofilm formation on the surface of these implants can cause recurrent infections and subsequent inflammatory responses in the host, resulting in tissue damage, necrosis, and re-hospitalization. To address these implant-associated infections, the best approach is to create antimicrobial coatings. Here, we report the fabrication of a biocompatible, non-leaching, and contact-based antibacterial coating for implants using quaternary pullulan functionalized MoS2 (MCP) glycosheets. The cationic MCP glycosheets were coated on the surfaces of polydopamine-modified stainless steel and polyvinyl fluoride substrates through a simple process of electrostatic interaction. The developed coating showed excellent antibacterial activity (>99.5%) against E. coli and S. aureus that remained stable over 30 days without leaching out of the substrates and retained its antibacterial activity. MCP-coated implants did not induce any acute or sub-chronic toxicity to mammalian cells, both in vitro and in vivo. Furthermore, MCP coating prevented S. aureus colonization on stainless steel implants in a mouse model of implant-associated infection. The MCP coating developed in this study represents a simple, safe, and effective antibacterial coating for preventing implant-associated infections.

Palladium Nanocapsules for Photothermal Therapy in the Near-Infrared II Biological Window.

Recent developments in nanomaterials with programmable optical responses and their capacity to modulate the photothermal effect induced by an extrinsic source of light have elevated plasmonic photothermal therapy (PPTT) to the status of a favored treatment for a variety of malignancies. However, the low penetration depth of near-infrared-I (NIR-I) lights and the need to expose the human body to a high laser power density in PPTT have restricted its clinical translation for cancer therapy. Most nanostructures reported to date exhibit limited performance due to (i) activity only in the NIR-I region, (ii) the use of intense laser, (iii) need of large concentration of nanomaterials, or (iv) prolonged exposure times to achieve the optimal hyperthermia state for cancer phototherapy. To overcome these shortcomings in plasmonic nanomaterials, we report a bimetallic palladium nanocapsule (Pd Ncap)─with a solid gold bead as its core and a thin, perforated palladium shell─with extinction both in the NIR-I as well as the NIR-II region for PPTT applications toward cancer therapy. The Pd Ncap demonstrated exceptional photothermal stability with a photothermal conversion efficiency of ∼49% at the NIR-II (1064 nm) wavelength region at a very low laser power density of 0.5 W/cm2. The nanocapsules were further surface-functionalized with Herceptin (Pd Ncap-Her) to target the breast cancer cell line SK-BR-3 and exploited for in vitro PPTT applications using NIR-II light. Pd Ncap-Her caused more than 98% cell death at a concentration of just 50 µg/mL and a laser power density of 0.5 W/cm2 with an output power of only 100 mW. Flow cytometric and microscopic analyses revealed that Pd Ncap-Her-induced apoptosis in the treated cancer cells during PPTT. Additionally, Pd Ncaps were found to have reactive oxygen species (ROS) scavenging ability, which can potentially reduce the damage to cells or tissues from ROS produced during PPTT. Also, Pd Ncap demonstrated excellent in vivo biocompatibility and was highly efficient in photothermally ablating tumors in mice. With a high photothermal conversion and killing efficiency at very low nanoparticle concentrations and laser power densities, the current nanostructure can operate as an effective phototherapeutic agent for the treatment of different cancers with ROS-protecting ability.

Quaternary Pullulan-Functionalized 2D MoS2 Glycosheets: A Potent Bactericidal Nanoplatform for Efficient Wound Disinfection and Healing.

Rapid emergence of multidrug-resistant bacterial strains has posed a global threat to public health. Hospital-acquired infections, especially in diabetic and burn patients, severely impede the process of wound healing, thereby causing high mortality. This calls for developing a new biomaterial that synergistically destroys pathogenic strains and also helps in promoting wound healing without causing any resistance generation. A new and highly potent antibacterial agent has been developed by integrating the bactericidal and wound healing properties of MoS2 nanosheets and a recently developed quaternized polysaccharide, pullulan (CP), into a single nanoplatform for accelerated wound therapy. MoS2 nanosheets are noncovalently functionalized with quaternized pullulan to yield glycosheets (MCP) that efficiently eradicate both Gram-negative Escherichia coli (5 µg/mL) and Gram-positive Staphylococcus aureus (10 µg/mL) within a short period of 4 h, through a synergistic action of membrane damage and chemical oxidation. MoS2 nanosheets coupled with CP exert a membrane-directed bactericidal action through distinct mechanisms of "pore-forming" and "non-pore-forming" pathways, respectively, whereas oxidative stress is induced by MoS2 nanosheets alone to collectively kill the pathogens. The MCP glycosheets have good biocompatibility and are also capable of disrupting and eradicating mature biofilms. Rapid and highly efficient in vivo wound disinfection and healing occurred upon MCP treatment through the reduction of inflammation and promotion of cellular proliferation and tissue remodeling. Thus, MCP glycosheets can emerge as a safe and potential biomaterial for better wound care management.

Formulation of a dual drug-loaded nanoparticulate co-delivery hydrogel system and its validation in rheumatoid arthritis animal model.

Rheumatoid arthritis (RA), a systemic autoimmune disease that dramatically affects patients' quality of life. Given the intricacy of RA's pathophysiology, no single treatment can completely halt the disease progression. Here, we attempted to treat RA holistically and synergistically by co-delivering methotrexate (MTX), a standard slow-acting anti-rheumatic drug, and phenethyl isothiocyanate (PEITC), a bioactive phytochemical, using a sodium alginate (SA)-pluronic F127 (PF-127) in situ hydrogel formulation. Therefore, in the current study, the co-delivery of MTX and PEITC in the nanoparticulate form could help enhance stability and solubility and facilitate greater penetration in the target arthritic tissues. The fabricated MTX NP and PEITC NE were found to have a minimum particle size, PDI, and good zeta potential. Results from in vitro release studies showed that MTX and PEITC were simultaneously released from the DD NP HG matrix over 6-7 days through diffusion and erosion mechanisms. An intra-articular (IA) injection of DD NP HG dramatically reduced chronic inflammation in adjuvant-induced arthritis (AIA) rats, delayed the onset of bone erosion, significantly reduced synovitis, and down-regulated the inflammatory cytokine expression. Most notably, the co-delivery strategy almost entirely restored the morphological features of the ankle joints of RA rats. The hepatic and renal function tests indicated good biological safety for DD NP HG in RA conditions. Taken together, these findings indicated that DD NP HG could achieve good anti-inflammatory activity and reverse cartilage disruption through a synergistic effect between two nanoparticulate forms of MTX and PEITC, which can effectively improve the drawbacks of their free forms.

A nanostructured dual-drug loaded smart hydrogel (DD NP HG) was successfully constructed for the intra-articular delivery of MTX and PEITC to the affected joints of RA.The fabrication of the nanoformulation of both MTX (MTX NP) and PEITC (PEITC NE) aided in mitigating the drawbacks and drug-related side effects of the free form of drugs.DD NP HG markedly suppressed joint inflammation and protect against bone destruction in arthritic rats.This combination approach of PEITC and MTX (DD NP HG) synergistically improved anti-arthritic activity and reduced the adverse side effects in vivo.

Thermosensitive smart hydrogel of PEITC ameliorates the therapeutic efficacy in rheumatoid arthritis.

Rheumatoid arthritis (RA) is an autoimmune condition that accompanies chronic inflammation of joints with limited therapeutic options. Phenethyl isothiocyanate (PEITC), a bioactive phytochemical, exerts its chemopreventive, anti-oxidant, and anti-inflammatory activity via the Nrf-2 pathway. However, limited water solubility, short half-life, and instability are reasons for the low bioavailability of PEITC that hampers clinical application. From studies in healthy rats, the performance of PEITC-loaded chitosan/pluronic F-127 smart hydrogel (PH) as a thermosensitive injectable demonstrated adequate thermosensitivity (gel formation), injectability (ease of administration), biocompatibility (with prolonged contact), pharmacokinetics (sustained drug release), and biosafety (nontoxic to major organs). In the adjuvant-induced arthritis (AIA) rat model, PEITC-hydrogel (PH50) injected into the knee joint lowered RA-related symptoms significantly (paw edema and arthritis score). Further, a marked reduction in bone erosion and inflammation-specific biomarkers was observed. Finally, this study demonstrates a smart injectable hydrogel optimally loaded with PEITC which is safe, biocompatible and exhibits significant therapeutic efficacy in RA conditions.

Quaternary ammonium substituted pullulan accelerates wound healing and disinfects Staphylococcus aureus infected wounds in mouse through an atypical 'non-pore forming' pathway of bacterial membrane disruption.

The emergence of multi-drug resistant pathogens has fueled the search for alternatives to the existing line of antibiotics that can eradicate pathogens without inducing resistance development. Here, we report the accelerated wound healing and disinfection potential of a non-amphiphilic quaternized fungal exopolysaccharide, pullulan, without resistance generation in pathogens. The quaternary ammonium substituted pullulan (CP) derivatives showed excellent bactericidal activity against both Gram negative (MBC90 = 1.5 µg mL-1) and Gram positive (MBC90 = 0.25 µg mL-1) bacteria at very low concentrations without showing any toxicity towards mammalian cells. A combined approach of atomistic molecular dynamics simulation and experimental assays revealed that CP exerts a membrane directed bactericidal action through an atypical "non-pore forming" pathway which is not yet established for any known antibacterial polysaccharides. This involves an increase in membrane roughness, disorder among anionic lipid tails, formation of localized anionic lipid clusters and membrane depolarization, finally leading to physical disruption of the membrane integrity. Moreover, CP also displayed biofilm eradication abilities and emerged as an excellent therapeutic material for disinfection and healing of infected wounds. The present work shows the potential of exploiting polysaccharides as next-generation broad-spectrum antimicrobials and provides a platform for further development of rationally designed pullulan-based functional materials for biomedical applications.

Zanthoxylum alatum ameliorates scopolamine-induced amnesia in rats: Behavioral, biochemical, and molecular evidence.

OBJECTIVE: Hydroethanolic extract of Zanthoxylum alatum seeds (HEZA) in scopolamine-induced amnesia was investigated for memory enhancing activity. MATERIALS AND METHODS: Radial arm maze (RAM) test was performed to evaluate the behavioral activity. Rats were treated with HEZA (50, 100, and 200 mg/kg, p. o.) and tacrine (3 mg/kg. i. p.) for 14 days. Scopolamine (0.4 mg/kg) was injected i. p. into rats after 45 min of drug administration on the 14th day. The messenger RNA (mRNA)/protein profile of few markers (acetylcholinesterase [AChE], heme oxygenase-1 [HO-1], nuclear factor-kappa B [NFκB], nuclear factor erythroid 2-related factor 2 [Nrf2], protein phosphatase 2A[PP2A], Tau, brain-derived neurotrophic factor [BDNF], tropomyosin-related kinase B [TrkB], Bcl-2-associated X protein [Bax], and Caspase-3) were also measured by polymerase chain reaction (PCR) and immunoblotting assay. Brain cytokines (tumor necrosis factor alpha [TNF-α], interleukin [IL]-1 ß, and IL-10) in hippocampus were evaluated using commercially available enzyme-linked immunosorbent assay kits. RESULTS: HEZA exhibited anti-amnesic activity as indicated by a significant reduction in the working memory error and reference memory error in RAM. Pretreatment with HEZA significantly down-regulated the expression of AChE, NFκB, Tau, Bax, and Caspase-3 with simultaneous up-regulation of Nrf2, HO-1, PP2A, BDNF, and TrkB genes in the hippocampal tissues similar to tacrine when compared with scopolamine-treated rats. Pretreatment with HEZA attenuated scopolamine-induced elevation of TNF-α, IL-1 ß, levels in hippocampus and reversed diminished IL-10 concentrations towards normal levels in the brain. CONCLUSION: Zanthoxylum alatum seeds could probably counteract amnesia. Since its use is mainly reported as a stimulant and tonic, this novel activity could be a boon for the scientists to explore more in this direction.

Zanthoxylum alatum abrogates lipopolysaccharide-induced depression-like behaviours in mice by modulating neuroinflammation and monoamine neurotransmitters in the hippocampus.

CONTEXT: Depression is an inflammatory, commonly occurring and lethal psychiatric disorder having high lifetime prevalence. Zanthoxylum alatum Roxb. (Rutaceae), commonly called Timur, has high medicinal value and is used ethnomedicinally for the treatment of various diseases. OBJECTIVE: To evaluate the effect of hexane extract of Z. alatum seeds (ZAHE) on lipopolysaccharide (LPS)-induced depression-like behaviour in Swiss albino mice. MATERIALS AND METHODS: Mice were treated with ZAHE (100 and 200 mg/kg, p.o.) and imipramine (10 mg/kg injected i.p.) for 14 days. On 14th day of the treatment, depression-like behaviour was induced by LPS (0.83 mg/kg injected i.p.) and after 24 h of LPS administration, it was assessed by measuring behavioural parameters and biochemical estimations. RESULTS: Behavioural tests, including the open field test, forced swimming test, tail suspension test and sucrose preference test revealed that ZAHE (100 and 200 mg/kg, p.o.) and imipramine (10 mg/kg injected i.p.) alleviated the depression symptoms of LPS-induced mice. Moreover, ZAHE treatments reversed the LPS-induced alterations in the concentrations of norepinephrine and serotonin (5-HT) and inhibited the expression of brain-derived neurotrophic factor, pro-inflammatory cytokines and oxido-nitrosative stress in the mice. Acute toxicity was calculated to be LD50 > 2500 mg/kg. DISCUSSION AND CONCLUSIONS: This study showed that LPS-induced depression in mice was significantly prevented by ZAHE at both the dosages. In conclusion, ZAHE exhibited an antidepressant activity by altering monoaminergic neurotransmitters in the brain combined with its anti-inflammatory potential. Thus, it could be an effective therapeutic against inflammation-induced depression and other brain disorders.

Role of Elsholtzia communis in counteracting stress by modulating expression of hspa14, C/EBP homologous protein, nuclear factor (erythroid-derived 2)-like-2 factor, Caspase-3, and brain-derived neurotrophic factor in rat hippocampus.

OBJECTIVE: Elsholtzia communis (Collett and Hemsl.) Diels has been widely distributed and is reported for many therapeutic effects. The present study aims to investigate the antistress activity of the leaf extract and its possible molecular mechanism. MATERIALS AND METHODS: Hydroethanolic extract of leaves of E. communis (100 and 200 mg/kg, p.o.) were administered for 7 days to stress-induced male Wistar rats. The experimental animals were divided into five groups (n = 6). The mRNA/protein profile of few stress responsive chaperones (hspa14), endoplasmic reticulum stress markers (C/EBP homologous protein [CHOP]), antioxidant regulating genes (nuclear factor (erythroid-derived 2)-like-2 factor [Nrf2]), apoptotic factors (Caspase-3) in rat hippocampus were studied by polymerase chain reaction and immunoblotting. RESULTS: The stress-related genes such as hspa14, CHOP, antioxidant gene Nrf2, apoptotic gene Caspase-3 which were overexpressed in the stress control group were significantly suppressed following administration of the extract at both the doses and the standard drug Ginseng. Likewise, brain-derived neurotrophic factor which is closely related with stress, was downregulated in the stress control group, was found to be upregulated following treatment with the extract and the standard drug Ginseng. CONCLUSION: Our findings clearly indicate that E. communis was able to counteract stress. Hence, it has the potential to develop as adaptogen and also as a replacement/substitute of the popularly used drug, Ginseng or Ashwagandha, which is on the verge of extinction or becoming endemic due to overuse.

Anticholinergic, antihistaminic, and antiserotonergic activity of n-hexane extract of Zanthoxylum alatum seeds on isolated tissue preparations: An ex vivo study.

OBJECTIVES: The aim of this study was to evaluate anticholinergic, antihistaminic, and antiserotonergic activity of the n-hexane extract of the seeds of Zanthoxylum alatum (ZAHE) on isolated ileum of rat and guinea pig and fundus of rat. MATERIALS AND METHODS: ZAHE was prepared using soxhlet extraction and cumulative concentration response curves were constructed using various doses on the tissues for acetylcholine (ACh), 5-hydroxytryptamine (5-HT), and histamine with or without n-hexane extract. Atropine, ketanserin, and pheniramine maleate were used as antagonists for ACh, serotonin, and histamine, respectively. RESULTS: ZAHE-induced concentration-dependent inhibition of isolated ileum and fundus in rat and ileum of guinea pig. The half maximal effective concentration (EC50) of ACh in the presence of atropine (10-6 M; P < 0.05) and ZAHE (1000 µg/ml; P < 0.01) was significantly higher than EC50of ACh alone. The EC50of 5-HT in the presence of ketanserin (10-5 M; P < 0.01) and ZAHE (1000 µg/ml; P < 0.05) was higher than EC50of 5-HT alone. Similarly, the EC50of histamine in the presence of pheniramine maleate (10-6 M; P < 0.01) and ZAHE (300 µg/ml; P < 0.01 and 1000 µg/ml; P < 0.05) was also significantly higher than EC50of histamine alone. CONCLUSION: From the study, it was observed that ZAHE shows significant anticholinergic, antiserotonergic, and antihistaminic activity. The study provides sufficient evidence that the seeds can be used in gastric disorders, cough, chest infection, etc., as per folklore claims.

Naringenin ameliorates inflammation and cell proliferation in benzo(a)pyrene induced pulmonary carcinogenesis by modulating CYP1A1, NFκB and PCNA expression.

Lung cancer is the major cause of cancer-related mortality and is a growing economic burden worldwide. Chemoprevention has emerged as a very effective preventive measure against carcinogenesis and several bioactive compounds in diet have shown their cancer curative potential on lung cancer. Naringenin (NRG), a predominant flavanone found in citrus fruits has been reported to possess anti-oxidative, anti-inflammatory and anti-proliferative activity in a wide variety of cancer. The aim of the present study is to divulge the chemopreventive nature of NRG against benzo(a)pyrene (B[a]P) induced lung carcinogenesis in Swiss albino mice. Administration of B[a]P (50mg/kg, p.o.) to mice resulted in increased lipid peroxidation (LPO), proinflammatory cytokines (TNF-α, IL-6 and IL-1ß) with subsequent decrease in activities of tissue enzymic antioxidants (SOD, CAT, GPx, GR, GST) and non-enzymic antioxidants (GSH and Vit-C). Treatment with NRG (50mg/kg body weight) significantly counteracted all these alterations thereby showing potent anti-cancer effect in lung cancer. Moreover, assessment of protein expression by immunoblotting and mRNA expression by RT-PCR revealed that NRG treatment effectively negates B[a]P-induced upregulated expression of CYP1A1, PCNA and NF-κB. Further, the antiproliferative effect of NRG was confirmed by histopathological analysis and PCNA immunostaining in B[a]P induced mice which showed increased PCNA expression that was restored upon NRG administration. Overall, these findings substantiate the chemopreventive potential of NRG against chemically induced lung cancer in mice.