Thymic and Peripheral T-cell Polarization in an Experimental Model of Russell's Viper Venom-induced Acute Kidney Injury.

Venom pathology is not restricted to the direct toxic effects of venom. Immunoinflammatory alteration as the etiology of snake venom-induced acute kidney injury (SAKI) is a less trodden path toward the development of alternative therapeutic approach. In the present study, we have associated the crest of renal damage stage to the immunological alteration, as reflected in thymic and peripheral T cell polarization in the murine model of SAKI. Renal injury in mice was confirmed from significant dysuresis and adversely altered biochemical renal markers. Histopathological alterations, as revealed by marked tubular and glomerular damage, reaffirmed kidney injury. SAKI is accompanied by significant inflammatory changes as indicated by neutrophilic leucocytosis, increased neutrophil to lymphocyte ratio and plasma CRP levels. Thymic immunophenotyping revealed significantly increased CD8+ cytotoxic T cell, and CD25+ both single positive population (p = .017-0.010) and CD44-CD25+ double negative population (DN3) (p = .002) accompanied by an insignificantly reduced CD4+ helper T cells (p = .451). Peripheral immunophenotyping revealed similar pattern as indicated by reduced helper T cells (p = .002) associated with significantly elevated cytotoxic T cells (p = .009) and CD25+ subset of both helper (p = .002) and cytotoxic (p = .024) T cells. The IL-10+ subset of both CD25+ and CD25- T cells were also found to be significantly elevated in the SAKI group (p ≤ 0.020) suggesting an immunosuppressive phenotype in SAKI. It can be concluded that T cells responds to venom-induced renal injury particularly through IL-10+ reparative phenotypes which are known for their immunosuppressive and anti-inflammatory activity.

Anti-inflammatory activity of cyclo-oxygenase2 inhibitory anionic protein fraction from Lamellidens marginalis (Lamarck).

Aqueous extract of freshwater mussel, Lamellidens marginalis is known to possess potent antioxidant and anti-inflammatory activity. Here, we have made an attempt to purify anti-inflammatory protein from Lamellidens marginalis extract (LME). Aqueous LME was prepared, and total protein was precipitated by 60% ammonium sulfate followed by purification through ion exchange chromatography. Isolated fractions were studied for anti-inflammatory activity in in vitro and in vivo experimental models. Active fractions were characterized by SDS PAGE and HPLC. Protein recovered from ammonium sulfate precipitation showed four distinct peaks in diethyl-aminoethyl cellulose ion exchange chromatography when eluted with stepwise salt gradient. Protein fraction eluted in 0.5 M sodium chloride solution showed maximum specific activity and anti-inflammatory activity in acute model and adjuvant induced chronic inflammation model. This fraction also showed cyclo-oxygenase 2 (COX2) enzyme inhibitory activity in in-vitro system. In SDS-PAGE 0.5 M NaCl fraction showed multiple bands after Coomassie brilliant blue staining and three distinct peaks in HPLC. In this study, we identified an anti-inflammatory protein fraction with high anionic property which could be attributed to inhibition of COX2 enzyme activity.

Prevention of arthritis markers in experimental animal and inflammation signalling in macrophage by Karanjin isolated from Pongamia pinnata seed extract.

Karanjin, the furanoflavonoid reported to possess gastroprotective and anti-diabetic properties, was investigated against experimental arthritis and its molecular signalling in inflammation was explored in macrophages. Karanjin was isolated from hexane extract of Pongamia pinnata seeds and was evaluated on arthritis markers in adjuvant induced arthritis model (AIA) in two doses (per oral; 10 mg/kg/day and 20 mg/kg/day). Karanjin dose dependently reduced collagen and cartilage breakdown markers viz. urinary hydroxyproline and glucosamine, respectively, serum lysosomal enzymes responsible for articular cartilage damage, and major proinflammatory cytokine TNFα, secreted by macrophages involved in articular inflammation and destruction. Karanjin also prevented joint damage as evidenced from arthritis score, radiographic and histopathological analysis. To delineate the molecular target of Karanjin, in vitro study on LPS induced macrophages were performed at calibrated non toxic doses (4 µg/mL and 6 µg/mL). Karanjin reduced TNFα production and also showed potent inhibitory effect on nitric oxide and reactive oxygen species production which is generally induced by TNFα from activated macrophages. NF-κB, the key regulator of TNFα signalling during inflammation was significantly suppressed by Karanjin. Our study for the first time highlights the anti-inflammatory role of Karanjin in experimental arthritis model as well as on macrophage signalling, thereby depicting its probable mechanism of action.

Antioxidant content and activity of the Indian fresh-water pearl mussel in the prevention of arthritis in an experimental animal model.

The flesh of the Indian fresh-water mussel, Lamellidens marginalis (LM; Lamarck, 1819), is the byproduct of pearl culture and a cheap protein source. The present study investigated the antioxidant content of this ethnomedicinally cited species to outline its importance in food security and disease prevention. LM was found to be rich in polyphenol antioxidants with good correlation with its reducing capacity. LM also showed a significant free-radical-scavenging activity, H(2)O(2)-scavenging activity and Fe-chelating activity. To study the effect of this dietary antioxidant against oxidative stress, we took inflammatory arthritis as a model. LM-treated arthritis rats showed a higher antioxidant defence system with elevated superoxide dismutase, total thiol, glutathione S transferase, glutathione peroxidase, total antioxidant status and catalase concentration of haemolysate. Oxidative stress markers like serum thiobarbituric acid-reacting substances, methyl glyoxal, NO and total oxidant status levels were decreased in LM-treated arthritis rats. Hence, the dietary antioxidants of LM were found to be effective in the prevention of oxidative stress in inflammatory arthritis. In conclusion, LM, the cash-crop byproduct, provides a rare opportunity for income and nutrition, not only by providing cheap and available energy, protein and dietary factors, but also by providing antioxidants effective against chronic inflammatory disease.

Anticancer potential of animal venoms and toxins.

Anticancer drug development from natural resources are ventured throughout the world. Animal venoms and toxins a potential bio resource and a therapeutic tool were known to man for centuries through folk and traditional knowledge. The biodiversity of venoms and toxins made it a unique source of leads and structural templates from which new therapeutic agents may be developed. Venoms of several animal species (snake, scorpion, toad, frog etc) and their active components (protein and non protein toxins, peptides, enzymes, etc) have shown therapeutic potential against cancer. In the present review, the anticancer potential of venoms and toxins from snakes, scorpions, toads and frogs has been discussed. Some of these molecules are in the clinical trials and may find their way towards anticancer drug development in the near future. The implications of combination therapy of natural products in cancer have been discussed.

Herbs and herbal constituents active against snake bite.

Snake bite, a major socio-medical problem of south east asian countries is still depending on the usage of antisera as the one and only source of treatment, which has its own limitations. In India, mostly in rural areas, health centres are inadequate and the snake bite victims mostly depend on traditional healers and herbal antidotes, as an alternative treatment. The present review has been focussed on the varied folk and traditional herbs and their antisnake venom compounds, which might be a stepping stone in establishing the future therapy against snake bite treatment and management.

Regulation of key molecules of immunological synapse by T11TS immunotherapy abrogates Cryptococcus neoformans infection in rats.

Cryptococcus neoformans infects and disseminates in hosts with diminished T cell responses. The immunomodulator T11TS (T11 target structure) had profound potential in glioma as well as C. neoformans infected model for disease amelioration. It is been established by our group that T11TS potentiates Calcineurin-NFAT pathway in T cells of C. neoformans infected rats. We investigated the upstream Immunological Synapse (IS) molecules that are vital for the foundation of initial signals for downstream signaling, differentiation and proliferation in T cells. Improved RANTES level in the T11TS treated groups suggests potential recruitment of T cells. Down-regulation of TCRαß, CD3ζ, CD2, CD45 and CD28 molecules by cryptococcus were boosted after T11TS therapy. Heightened expression of inhibitory molecule CTLA-4 in cryptococcosis was dampened by T11TS. The decline of MHC I, MHC II and CD80 expression on macrophages by C. neoformans were enhanced by T11TS. The dampening of positive regulators and upsurge of negative regulators of the IS during cryptococcosis was reversed with T11TS therapy resulting in enhanced clearance of fungus from the lungs as envisaged by our histological studies. This preclinical study with T11TS opens a new prospect for potential immunotherapeutic intervention against the devastating C. neoformans infection with positive aspect for the long-term solution and a safer immunotherapeutic regimen.

Experimental osteoporosis induced in female albino rats and its antagonism by Indian black scorpion (Heterometrus bengalensis C.L.Koch) venom.

The present study was designed to explore the antiosteoporosis activity of the Indian black scorpion (Heterometrus bengalensis) venom on experimental osteoporosis female albino rats. Sham operated control rats were designated as Gr I, Gr II animals served as osteoporosis control, Gr III osteoporosis rats were treated with SV (1/25th of MLD), Gr IV osteoporosis rats were treated with 1/50th of MLD of SV and Gr V osteoporosis rats were treated with standard (calcium and vit-D3). As compared with the Gr I rats, the Gr II rats showed typical osteoporosis changes in increased of urinary Ca(2+), PO(4)(3-), CRE, OH-P levels, serum/plasma Ca(2+), PO(4)(3-), TRAP, IL1, IL6, TNFalpha and PTH level, bone Ca(2+), PO(4)(3-), Mg(2+), Zn(2+) and decreased level of serum/plasma ALP, EST and PTH, bone Na(+). In Gr III, Gr IV and Gr V rats, the osteoporosis changes of urine, serum and bone, were significantly restored as compared with the Gr II rats. The bone dimensions, morphology and histological changes observed in Gr II rats were restored in Gr III, Gr IV and Gr V rats. This study confirms that the Indian black scorpion venom may influence bone remodeling process by stimulating bone formation and reducing bone resorption process of osteogenesis.

Effect of cellulose nanocrystals on the performance of drug loaded in situ gelling thermo-responsive ophthalmic formulations.

Triblock poloxamer copolymer (PM) has been extensively utilized to deliver various ophthalmic pharmaceutical compounds. The aim of efficient ophthalmic drug delivery strategy is to attain the longer precorneal resident time and good bioavailability of drugs. In this pursuit, the influence of cellulose nanocrystals (CNC) on the in situ gelation behavior of PM and in vitro release of pilocarpine hydrochloride from the nanocomposites formulations was studied. The critical concentration of gelation of PM being 18% (wt/v) was dropped to 16.6% (wt/v) by the addition of a very low percentage of CNC. The reinforcing nature of CNC via H-bonding in the in situ nanocomposite gel also led to an increase in gel strength along with the sustained release of loaded drugs when compared with the pure PM gel. All formulations revealed that the drug release mechanism is controlled by the Fickian diffusion. Thus, the CNC has a significant effect on the gelation behavior, gel strength, and drug release kinetics of PM-CNC formulations.

Synthesis of methylcellulose/cellulose nano-crystals nanocomposites: Material properties and study of sustained release of ketorolac tromethamine.

Non-toxic nanocomposites based bio-films obtained from methylcellulose (MC) can reduce environmental problems associated with synthetic polymers. A new facile route for the isolation of cellulose nano-crystals (CNC) from jute waste is successfully utilized here. The fabrication of CNC reinforced MC nanocomposites by film casting technique and the studies of the effect of CNC on the properties of the MC based nanocomposites have been reported. The synthesized nanocomposites have shown improved UV resistance, mechanical, barrier, and thermal properties. FTIR results established the physicochemical compatibility between the drug, MC and CNC in nanocomposites. In vitro permeation studies performed by using Franz diffusion cell revealed diffusion mediated sustained drug release from the devices due to the presence of interaction between MC and CNC through H-bonding, electrostatic interaction between the hydrophilic polymer/CNC chains with the drug and the formation of tortuous path. The nanocomposites can be used for edible packaging and transdermal drug delivery.

Development of active packaging material based on cellulose acetate butyrate/polyethylene glycol/aryl ammonium cation modified clay.

Active packaging is one of the interesting concepts in food industry which extend the shelf-life of the food products. The purpose of this work was to develop nontoxic antimicrobial nanocomposite films. Benzyltrimethylammonium chloride modified montmorillonite (BMMT) were used as nano-filler and the prepared BMMT was characterized using Fourier transform infrared spectroscopy (FTIR), X-ray diffraction (XRD), and thermogravimetric analysis. Polyethylene glycol (PEG) plasticized cellulose acetate butyrate (CAB) films with different ratios of PEG and CAB was prepared and it was found that the 20 wt% PEG in CAB matrix (CBP20) gave optimal results in terms of mechanical properties. BMMT was mixed with CBP20 in different proportions to prepare nanocomposites. 3 wt% BMMT loaded nanocomposite gave best in terms of the barrier and mechanical properties. The storage modulus, thermal stability, glass transition, and melting temperature of the nanocomposites increased with the loading of 1, 3, and 5 wt% of BMMT. Furthermore, these nanocomposites showed nontoxic and antimicrobial behavior.

Hematological alterations and splenic T lymphocyte polarization at the crest of snake venom induced acute kidney injury in adult male mice.

Snake venom induced acute kidney injury (SAKI) is of great clinical relevance in tropical countries. Involvement of T cell, a key mediator of AKI and its remission, is least explored in SAKI. In the present study the in vivo hematological alterations and associated splenic T cell polarization is probed in order to investigate the immune response at the crest of Russell's viper venom (RVV) induced AKI in experimental murine model. Based on a dose and time kinetic study intra muscular injection dose of 20 µg RVV/100 gm body weight of mice and incubation period of 60 h was selected for induction of SAKI. Renal involvement in SAKI group was confirmed from oliguria, significantly elevated urinary microprotein (p < 0.001), decreased urinary creatinine (p = 0.003) and creatinine clearance (p < 0.001) compared to control. Hematological analyses revealed a significant neutrophilic leukocytosis (p < 0.001) associated with a reduced lymphocyte percentage (p < 0.001) favoring a state of acute inflammation in SAKI group. Immunophenotyping study of splenocytes showed a significant decrease in CD4+/CD8+ ratio (p < 0.001) with a significant increase in regulatory (CD25+FoxP3+) helper and cytotoxic subset of T cell (p < 0.001). Significant increase in IL-10+ regulatory helper and cytotoxic T cell (p < 0.001) further confirmed the internal milieu favoring immunosuppression. Apart from these the CD25-FoxP3+ reservoir regulatory T cells were also found to be significantly elevated in SAKI group compared to that of control (p < 0.001). Taken together, the results of the present study clearly indicated a state of acute inflammation and splenic T cell polarization towards regulatory subset at the crest of SAKI.

pH-sensitive chitosan/alginate core-shell nanoparticles for efficient and safe oral insulin delivery.

Chitosan-alginate (CS/ALG) nanoparticles were prepared by formation of an ionotropic pre-gelation of an alginate (ALG) core entrapping insulin, followed by chitosan (CS) polyelectrolyte complexation, for successful oral insulin administration. Mild preparation process without harsh chemicals is aimed at improving insulin bio-efficiency in in vivo model. The nanoparticles showed an average particle size of 100-200 nm in dynamic light scattering (DLS), with almost spherical or sub-spherical shape and ∼ 85% of insulin encapsulation. Again, retention of almost entire amount of encapsulated insulin in simulated gastric buffer followed by its sustained release in simulated intestinal condition proved its pH sensitivity in in vitro release studies. Significant hypoglycemic effects with improved insulin-relative bioavailability (∼ 8.11%) in in vivo model revealed the efficacy of these core-shell nanoparticles of CS/ALG as an oral insulin carrier. No systemic toxicity was found after its peroral treatment, suggesting these core-shell nanoparticles as a promising device for potential oral insulin delivery.

Quantifying morphological alteration of RBC population from light scattering data.

Studies of RBC morphological alterations, despite their potential clinical and experimental application, are compromised due to lack of simple and rapid techniques. As a complementary approach toward quantitative microscopy, we have reconstituted morphological information from light scattering data obtained from flow cytometer. Normal and poikilocytic agent treated samples were analyzed by microscopy and respective morphological index (MI) was calculated from the morphology based scores assigned to RBC. The samples were simultaneously analyzed by flowcytometer and the scatter data were obtained. Accordingly, the best correlated parameters of both forward scatter and side scatter were chosen to formulate a suitable regression model with MI as response. Flow cytometry data was also verified with another instrument (BD FACS Verse) and the equation obtained was validated with separate set of samples. The multivariate regression analysis yields a quadratic model with MI as response (R2 = 0.96, p <  0.001). The flow cytometric data from both instruments were in good agreement (Intra class correlation ∼0.9, p <  0.001). The model was found to simulate the sample MI with high accuracy (R2 = 0.97, p <  0.001). This proposed method was verified to be simple, rapid, quantitative and cost effective for the measurement of morphological alteration of RBC.

Viper and cobra venom neutralization by alginate coated multicomponent polyvalent antivenom administered by the oral route.

BACKGROUND: Snake bite causes greater mortality than most of the other neglected tropical diseases. Snake antivenom, although effective in minimizing mortality in developed countries, is not equally so in developing countries due to its poor availability in remote snake infested areas as, and when, required. An alternative approach in this direction could be taken by making orally deliverable polyvalent antivenom formulation, preferably under a globally integrated strategy, for using it as a first aid during transit time from remote trauma sites to hospitals. METHODOLOGY/PRINCIPAL FINDINGS: To address this problem, multiple components of polyvalent antivenom were entrapped in alginate. Structural analysis, scanning electron microscopy, entrapment efficiency, loading capacity, swelling study, in vitro pH sensitive release, acid digestion, mucoadhesive property and venom neutralization were studied in in vitro and in vivo models. Results showed that alginate retained its mucoadhesive, acid protective and pH sensitive swelling property after entrapping antivenom. After pH dependent release from alginate beads, antivenom (ASVS) significantly neutralized phospholipaseA2 activity, hemolysis, lactate dehydrogenase activity and lethality of venom. In ex vivo mice intestinal preparation, ASVS was absorbed significantly through the intestine and it inhibited venom lethality which indicated that all the components of antivenom required for neutralization of venom lethality were retained despite absorption across the intestinal layer. Results from in vivo studies indicated that orally delivered ASVS can significantly neutralize venom effects, depicted by protection against lethality, decreased hemotoxicity and renal toxicity caused by russell viper venom. CONCLUSIONS/SIGNIFICANCE: Alginate was effective in entrapping all the structural components of ASVS, which on release and intestinal absorption effectively reconstituted the function of antivenom in neutralizing viper and cobra venom. Further research in this direction can strategize to counter such dilemma in snake bite management by promoting control release and oral antivenom rendered as a first aid.

pH sensitive N-succinyl chitosan grafted polyacrylamide hydrogel for oral insulin delivery.

pH sensitive PAA/S-chitosan hydrogel was prepared using ammonium persulfate (APS) as an initiator and methylenebisacrylamide (MBA) as a crosslinker for oral insulin delivery. The synthesized copolymer was characterized by Fourier transform infrared spectroscopy (FT-IR) and X-ray diffraction (XRD) study; morphology was observed under scanning electron microscope (SEM). The PAA/S-chitosan with ∼ 38% of insulin loading efficiency (LE) and ∼ 76% of insulin encapsulation efficiency (EE), showed excellent pH sensitivity, retaining ∼ 26% of encapsulated insulin in acidic stomach pH 1.2 and releasing of ∼ 98% of insulin in the intestine (pH 7.4), providing a prolonged attachment with the intestinal tissue. The oral administration of insulin loaded PAA/S-chitosan hydrogel was successful in lowering the blood glucose level of diabetic mice. The bioavailability of insulin was ∼ 4.43%. Furthermore, no lethality or toxicity was documented after its peroral administration. Thus, PAA/S-chitosan hydrogel could serve as a promising oral insulin carrier in future.

Suppression of NF-κB p65 nuclear translocation and tumor necrosis factor-α by Pongamia pinnata seed extract in adjuvant-induced arthritis.

Pongamia pinnata is a plant known for its therapeutic usage in Indian traditional medicine. Despite the controversy regarding toxic flavonoid and erucic acid content, the seed of this plant is consumed in tribal medicine and its oil is used in Ayurveda to treat psoriasis and arthritis. This study explored the potential anti-arthritic effects of a P. pinnata seed (hexane) extract (PSE) at non-lethal doses in an adjuvant-induced arthritic rat model; possible mechanisms of any observed effects were also explored. After establishing the lethal doses arising from oral exposure to the extract, the material was administered per os daily at two doses (0.3 g/kg/day; 0.5 g/kg/day) to arthritic rats. Other rats received indomethacin or vehicle (control). Treatments were performed for a total of 14 days. One day after the final exposure, the rats were euthanized to permit harvest of various cells, blood, and tissues for analyses. Paw diameter and tissue myeloperoxidase activity in the paws were evaluated as indices for edema and neutrophil infiltration into the tissue. The severity of arthritis in the experimental rats was assessed via measures of urinary hydroxyproline (HP) and glucosamine, and of serum pro-inflammatory TNFα and anti-inflammatory IL-10. The extent of NF-κB p65 nuclear translocation in peritoneal macrophages harvested from naïve rats and then treated in vitro was also assessed. The results indicated that exposure to PSE significantly decreased paw diameter, tissue myeloperoxidase level, and levels of urinary HP and glucosamine, as well as of serum TNFα and IL-10 in adjuvant-injected (arthritic) rats. In vitro PSE treatment also resulted in a marked inhibition of NF-κB p65 nuclear translocation in primary cultures of peritoneal macrophages. Thus, PSE appears to be able to prevent experimental arthritis, in part, by helping to maintain the balance between pro- and anti-inflammatory cytokines and by inhibiting NF-κB activation.

Indian freshwater edible snail Bellamya bengalensis lipid extract prevents T cell mediated hypersensitivity and inhibits LPS induced macrophage activation.

ETHNOPHARMACOLOGICAL RELEVANCE: Soup prepared from the foot of fresh water edible snail, Bellamya bengalensis, is traditionally consumed by the tribes of Jharkhand against rheumatism like bone and joint inflammation. As rheumatism has underlying involvement of cell mediated hypersensitivity, in vivo delayed-type hypersensitivity (DTH) model and in vitro LPS-induced macrophage signaling were studied to delineate the mechanism by which Bellamya bengalensis exerts its ethnomedicinal function. Since the whole meat is consumed, the lipid of Bellamya bengalensis (BBL) was hypothesized to be the active part. METHODS AND MATERIALS: BBL isolated from the foot part of this species, was characterized and given by gavage daily (10mg BBL/kg; 20mg BBL/kg) to mice for 3 weeks prior to initiating development of DTH. Effects of DTH induced changes in paw diameter, serum nitric oxide (NO), serum tumor necrosis factor (TNF)-α level, CINC1 level, splenic CD4(+)/CD8(+) cell ratios, and level of splenic Treg cells were then compared with values in untreated control mice. In vitro effect of BBL on LPS-stimulated macrophage, the immune cell that is active in DTH, was assessed by NF-kB p65 nuclear translocation, reactive oxygen species (ROS), TNFα, and NO production. RESULTS: BBL was characterized, and its supplementation in situ led to significant decrease in paw edema, tissue myeloperoxidase activity, NO level, serum TNFα level and CINC 1 level as well as decrease in splenic CD4(+)/CD8(+) ratios and increase in level of Treg cells. BBL was shown to inhibit ROS, NO, and TNFα production along with NF-kB p65 nuclear translocation in LPS stimulated macrophage. CONCLUSION: Bellamya bengalensis, traditionally used against diseases with underlying etiology of cell mediated immunity as in rheumatism, which acts through inhibition of overexpressed cell mediated immunity. The factor exerting this activity probably is the oleic acid and cyclopropane fatty acid rich lipid, isolated after the ethnomedicinal clue, from the foot of this species.

Taro corms mucilage/HPMC based transdermal patch: an efficient device for delivery of diltiazem hydrochloride.

The aim of this work is to examine the effectiveness of mucilage/hydroxypropylmethylcellulose (HPMC) based transdermal patch (matrix type) as a drug delivery device. We have successfully extracted mucilage from Colocasia esculenta (Taro) corms and prepared diltiazem hydrochloride incorporated mucilage/HPMC based transdermal patches using various wt% of mucilage by the solvent evaporation technique. Characterization of both mucilage and transdermal patches has been done by several techniques such as Molisch's test, organoleptic evaluation of mucilage, mechanical, morphological and thermal analysis of transdermal patches. Skin irritation test is studied on hairless Albino rat skin showing that transdermal patches are apparently free of potentially hazardous skin irritation. Fourier transform infrared analysis shows that there is no interaction between drug, mucilage and HPMC while scanning electron microscopy shows the surface morphology of transdermal patches. In vitro drug release time of mucilage-HPMC based transdermal patches is prolonged with increasing mucilage concentration in the formulation.

Oral insulin delivery by self-assembled chitosan nanoparticles: in vitro and in vivo studies in diabetic animal model.

We have developed self-assembled chitosan/insulin nanoparticles for successful oral insulin delivery. The main purpose of our study is to prepare chitosan/insulin nanoparticles by self-assembly method, to characterize them and to evaluate their efficiency in vivo diabetic model. The size and morphology of the nanoparticles were analyzed by dynamic light scattering (DLS), atomic force microscopy (AFM) and scanning electron microscopy (SEM). The average particle size ranged from 200 to 550 nm, with almost spherical or sub spherical shape. An average insulin encapsulation within the nanoparticles was ~85%. In vitro release study showed that the nanoparticles were also efficient in retaining good amount of insulin in simulated gastric condition, while significant amount of insulin release was noticed in simulated intestinal condition. The oral administrations of chitosan/insulin nanoparticles were effective in lowering the blood glucose level of alloxan-induced diabetic mice. Thus, self-assembled chitosan/insulin nanoparticles show promising effects as potential insulin carrier system in animal models.