Andrographolide-Soya-L-α-Phosphatidyl Choline Complex Augmented Solubility and Drug Delivery in Leishmania donovani, a Causative Agent for Cutaneous and Visceral Leishmaniasis.

The utility of andrographolide (AN) in visceral leishmaniasis (VL) and cutaneous leishmaniasis (CL) is limited owing to poor solubility, hindered permeation, and unstable structure under physiological conditions. The present study mainly focuses on synthesizing of andrographolide-Soya-L-α-phosphatidyl choline (ANSPC) complex in ethanol and its characterization using various spectral and analytical techniques. Results from FT-IR, 1H NMR, ROSEY, and in silico docking techniques suggest ANSPC complex formation due to inter-molecular interaction between the hydrophilic head of SPC and hydroxyl group of AN present at 24th position. ANSPC complex demonstrated the solubility of 113.93 ± 6.66 µg/mL significantly (P < 0.05) greater than 6.39 ± 0.47 µg/mL of AN. The particle size of ANSPC complex was found to be 182.2 ± 2.69 nm. The IC50 value of AN suspension (PBS, pH ~ 7.4) at 24, 48, and 72 h against Leishmania donovani (L. donovani) was noticed to be 32.76 ± 4.53, 20.87 ± 2.37, and 17.71 ± 3.06 µM/mL, respectively. Moreover, augmented aqueous solubility of ANSPC complex led to significant (P < 0.05) reduction in IC50 value, i.e., 25.02 ± 4.35, 11.31 ± 0.60, and 8.33 ± 2.71 µM/mL at 24, 48, and 72 h, respectively. The IC50 values for miltefosine were noted to be 9.84 ± 2.65, 12.13 ± 7.26, and 6.56 ± 0.61 µM/mL at similar time periods. Moreover, ANSPC complex demonstrated augmented cellular uptake at 24 h as compared to 6 h in L. donovani. We suppose that submicron size and phospholipid-mediated complexation might have endorsed the permeation of ANSPC complex across the plasma membrane of L. donovani parasite by transport mechanisms such as P-type ATPase. ANSPC complex warrants further in-depth in vivo studies under a set of stringent parameters for translating the product into a clinically viable form.

Applications of lipid-engineered nanoplatforms in the delivery of various cancer therapeutics to surmount breast cancer.

Breast cancer (BC) is the most extensively accounted malignancy among the women across the globe and is treatable in 70-80% of patients with early-stage, non-metastatic cancer. The current available therapies have been found to be less effective to treat distant organ metastases and advanced breast cancers. The clinical efficacy hugely suffers from chemoresistance, non-specific toxicity, relapse and other associated adverse effects. Furthermore, lack of controlled delivery and effective temporospatial presence of chemotherapeutics has resulted in suboptimal therapeutic response. Nanotechnology based approaches have been widely used over the period as they are nanometric, offer controlled and site-specific drug release along with reduced toxicity, improved half-life, and stability. Lipid-based nanoplatforms have grabbed a tremendous attention for delivering cancer therapeutics as they are cost-effective, scalable and provide better entrapment efficiency. In this review, all the promising applications of lipid-engineered nanotechnological tools for breast cancer will be summarized and discussed. Subsequently, BC therapy achieved with the aid of chemotherapeutics, phytomedicine, genes, peptides, photosensitizers, diagnostic and immunogenic agents etc. will be reviewed and discussed. This review gives tabular information on all the results obtained pertaining to the physicochemical properties of the lipidic nanocarrier, in vitro studies conferring to mechanistic drug release profile, cell viability, cellular apoptosis and in vivo studies referring to cellular internalisation, reduction of tumor volume, PK-PD profile, bioavailability achieved and anti-tumor activity in detail. It also gives complete information on the most relevant clinical trials done on lipidic nanoplatforms over two decades in tabular form. The review highlights the current status and future prospects of lipidic nanoplatforms with streamlined focus on cancer nanotherapeutics.

Nutritional and physiological responses of broiler chicken to the dietary supplementation of Moringa oleifera aqueous leaf extract and ascorbic acid in tropics.

This study was intended to determine the comparative effects of Moringa oleifera aqueous leaf extract (MOALE) and ascorbic acid supplementation in the drinking water on growth performance, haemato-biochemical profile, antioxidant status and immune response of broiler chickens under tropical climate. All 135-day-old broiler chicks were divided into 3 different treatment groups. T0 served as control, T1 were fed basal ration with MOALE (90 ml/L drinking water), and T2 offered basal ration with ascorbic acid (15 mg/L drinking water) for 35-day experiment. All the standard managemental practices were followed during the experimental period. The 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity of MOALE was calculated with reference to different ascorbic acid concentration as standard. The antioxidant activity percentage of MOALE (90 µl) was 93.89%, and ascorbic acid (15 µg) was 98.76%, respectively, which was nearer to cent percent. Therefore, 90 ml MOALE and 15 mg ascorbic acid per litre drinking water, respectively, were used for supplementation in broiler chicken. The growth performance and feed efficiency were better in MOALE group followed by ascorbic acid supplemented birds without affecting the metabolism of nutrients. However, 10.71% better FCR was noted in MOALE supplemented group followed by ascorbic acid group (6.28%) in comparison with control. Most of the haemato-biochemical profiles were unaffected by the treatment except creatinine, while antioxidant profile was improved in the treatment group. Immunity status of broiler chicken against NDV was enhanced in both treatment groups; however, maximum profit is obtained in MOALE group followed by ascorbic acid supplemented birds.

Pharmacophore modeling and molecular dynamics approach to identify putative DNA Gyrase B inhibitors for resistant tuberculosis.

One of the major mechanisms followed by the therapeutic agents to target the causative organism of TB, mycobacterium tuberculosis (Mtb), involves disruption of the replication cycle of the pathogen DNA. The process involves two steps that occur simultaneously, ie, breakage and reunion of DNA at gyrase A (GyrA) domain and ATP hydrolysis at gyrase B (GyrB) domain. Current therapy for multi-drug resistant TB involves FDA approved, Fluoroquinolone-based antibiotics, which act by targeting the replication process at GyrA domain. However, resistance against fluoroquinolones due to mutations in the GyrA domain has limited the use of this therapy and shifted the focus of the research community on the GyrB domain. Thus, this study involves in silico designing of chemotherapeutic agents for resistant TB by targeting GyrB domain. In the current study, a pharmacophore model for GyrB domain was generated using reported inhibitors. It was utilized as a query search against three commercial databases to identify GyrB domain inhibitors. Additionally, a qualitative Hip-Hop pharmacophore model for GyrA was also developed on the basis of some marketed fluoroquinolone-based GyrA inhibitors, to remove non-selective gyrase inhibitors obtained in virtual screening. Further, molecular dynamic simulations were carried out to determine the stability of the obtained molecules in complex with both the domains. Finally, Molecular mechanics with generalized Born and surface area solvation score was calculated to determine the binding affinity of obtained molecule with both domains to determine the selectivity of the obtained molecules that resulted in seven putative specific inhibitors of GyrB domain.

Counting on Fragment Based Drug Design Approach for Drug Discovery.

Fragment based drug design (FBDD) is a structure guided ligand design approach used in the process of drug discovery. It involves identification of low molecular weight fragments as hits followed by determination of their binding mode using X-ray crystallography and/or NMR spectroscopy. X-ray protein crystallography is one of the most sensitive biophysical methods used for screening and is least prone to false positives. It also provides detailed structural information of the protein-fragment complex at the atomic level. The retrieved binding information facilitates the optimization of fragments into drug like molecules. These identified molecules bind efficiently with the target proteins and form high quality binding interactions. Fragment-based screening using X-ray crystallography is, therefore, an efficient method for identifying binding hotspots on proteins that can be further exploited by chemists and biologists for the discovery of new drugs. The recent advancements in FBDD technique are illustrated in this review along with recently published success stories of FBDD technique in drug discovery.

Synergistic anti-glioma effect of a coloaded nano-drug delivery system.

The anti-glioma effect of temozolomide (Tem) is sometimes undermined by the emerging resistance. Recently, resveratrol (Res), herbal medicine extracted from grape seeds, has been demonstrated for its potential use in chemosensitization. In the current study, both these drugs were loaded simultaneously into nanoparticles with methoxy poly(ethylene glycol)-poly epsilon caprolactone (mPEG-PCL) as drug carriers in order to achieve better antitumor efficiency. Tem/Res-coloaded mPEG-PCL nanoparticles were constructed, characterized, and tested for antitumor effect on glioma cells by using in vitro and xenograft model system. The nanoparticle constructs were satisfactory with drug loading content (Res =~12.4%; Tem =~9.3%) and encapsulation capacity of >85% for both the drugs. In addition, the coencapsulation led to better in vitro stability of the nanoparticles than Tem-loaded nanoparticles. An in vitro uptake study demonstrated a high uptake efficiency of the nanoparticles by glioma cells. The synergistic antitumor effect against glioma cells was observed in the combinational treatment of Res and Tem. Tem/Res-coloaded nanoparticles induced higher apoptosis in U87 glioma cells as compared to cells treated by the combination of free drugs. Tem/Res-coloaded particles caused more effective inhibition of phosphor-Akt, leading to upregulation of the downstream apoptotic proteins. In addition, the in vivo study showed the superior tumor delaying effect of coloaded nanoparticles than that of free drug combination. These results suggest that Tem/Res-coloaded nanoparticles could be a potential useful chemotherapeutic formulation for glioma therapy.

Toxicophore exploration as a screening technology for drug design and discovery: techniques, scope and limitations.

Toxicity is a common drawback of newly designed chemotherapeutic agents. With the exception of pharmacophore-induced toxicity (lack of selectivity at higher concentrations of a drug), the toxicity due to chemotherapeutic agents is based on the toxicophore moiety present in the drug. To date, methodologies implemented to determine toxicophores may be broadly classified into biological, bioanalytical and computational approaches. The biological approach involves analysis of bioactivated metabolites, whereas the computational approach involves a QSAR-based method, mapping techniques, an inverse docking technique and a few toxicophore identification/estimation tools. Being one of the major steps in drug discovery process, toxicophore identification has proven to be an essential screening step in drug design and development. The paper is first of its kind, attempting to cover and compare different methodologies employed in predicting and determining toxicophores with an emphasis on their scope and limitations. Such information may prove vital in the appropriate selection of methodology and can be used as screening technology by researchers to discover the toxicophoric potentials of their designed and synthesized moieties. Additionally, it can be utilized in the manipulation of molecules containing toxicophores in such a manner that their toxicities might be eliminated or removed.

Histological changes in thalamus in short term survivors following traumatic brain injury: an autopsy study.

BACKGROUND: Severe traumatic brain injury (TBI) is a major cause of morbidity and mortality. Reduction of thalamic volumes were seen in upto 80% of patients who survived for more than 3 months after TBI. However, the same may not be true in patients who died earlier following TBI. AIMS: To to study the thalamus for evidence of any injury in short term survivors of TBI (<5 days) using immunohistochemistry to look for evidence of acute thalamic injury. MATERIALS AND METHODS: A cross sectional prospective study was done in which autopsy specimens of short term survivors of TBI (<5 days) were studied for histopathological changes. RESULTS: A total of 16 patients with a mean age of 37.8 years were included in the study. CT scan revealed acute subdural haematoma in 10, contusions in 4 patients, extradural haematoma and depressed fracture in 1 each, and diffuse axonal injury in 1 patient. Seven patients required surgery in the form of a decompressive hemicraniectomy. The histopathological analysis of the bilateral thalami showed evidence of congestion of the cerebral capillaries in 8 patients. Axonal retraction balls were seen in 8 patients, myelin breakdown products were seen in 14 patients and axonal swelling was seen in 14 patients. CONCLUSIONS: Thalamic injury is universal in the setting of severe TBI in patients who have decreased survival and may be a significant factor for the poor outcome in these patients.

Role of Apoptotic Proteins in REC-2006 Mediated Radiation Protection in Hepatoma Cell Lines.

The present study was carried out to evaluate the role of apoptotic proteins in REC-2006-mediated radiation protection in hepatoma cell lines. REC-2006 treatment 2 h before irradiation strongly inhibited the cleavage of ATM and PARP-1 in HepG2 cells. The expression of nuclear apoptosis inducing factor (AIF) was found to be more inhibited (~17%) in HepG2 cells in REC-2006 + radiation-treated group. More inhibition (~33%) of cytochrome c was observed in HepG2 cells upon REC-2006 treatment 2 h prior irradiation. Similarly, significantly more (P<.05) inhibition of Apaf-1, caspase-9 and caspase-3 was observed in REC-2006 + radition-treated group in HepG2 cells. REC-2006 treatment restored the expression of ICAD in HepG2 cells; however, no restoration was observed in Hep3B cells. Lower nuclear to cytoplasmic CAD ratio was observed in HepG2 cells (~0.6) as compared with Hep3B cells (~1.2) in REC-2006 + radiation-treated group. In conclusion, REC-2006 rendered higher protection in HepG2 cells by inhibiting the expression and translocation of AIF, inhibiting the cleavage of ATM and PARP-1, restoring the expression of ICAD, inhibiting the release of cytochrome c and thus modulating the expression of Apaf-1 caspase-9 and activity of caspase-3.

Delayed bilateral thalamic bleeding post-ventriculoperitoneal shunt.

Ventriculoperitoneal shunt is one of the most common procedure done by neurosurgeon worldwide. We present a rare case of delayed intracerebral bleed post VP shunt and discuss the possible causes of intracerebral hemorrhage post ventriculoperitoneal shunt and bilateral thalamic bleed. The presumed cause in our case was shunt induced disseminated coagulation profile.

Autologous transfusion of drain contents in elective primary knee arthroplasty: its value and relevance.

BACKGROUND: Total knee arthroplasty is associated with significant post-operative blood loss often necessitating blood transfusions. Blood transfusions may be associated with transfusion reactions and may transmit human immunodeficiency virus, hepatitis C virus and hepatitis B virus, with devastating consequences. After total knee arthroplasty, transfusion of the contents of an autologous drain is becoming common practice. The aim of our study was to look at the effectiveness of these drains in elective primary total knee arthroplasty. MATERIALS AND METHODS: A prospective study was conducted including 70 non-randomised patients. A normal suction drain was used in 35 patients (group A), whereas in the other 35 patients, a CellTrans™ drain was used (group B). All the operations were performed by four surgeons using a tourniquet with a medial parapatellar approach. Pre- and post-operative haemoglobin concentrations were recorded in both groups. A Student's t-test was applied to determine the statistical significance of the data collected. RESULTS: The average fall in post-operative haemoglobin was 3.66 g/dL (SD 1.46; range, 0.6-7.0) among patients in whom the simple drain was used (group A) and 2.29 g/dL (SD 0.92; range, 0.6-5.9) among those in whom the CellTrans™ drain was used (group B) (p<0.0001). Twenty-five units of allogeneic blood were required in group A compared to four units in group B. The rate of transfusion was 5.7% (2 patients) in the group in which CellTrans™ drain was used and 25.7% (9 patients) in the group in which a simple suction drain was used. DISCUSSION: Total knee arthroplasty is associated with significant post-operative blood loss despite best operative technique. Autologous reinfusion of the contents of a CellTrans™ drain significantly reduces the rate of post-operative blood transfusion. This study indicates that the use of an autologous drain could be recommended as routine practice in primary total knee arthroplasty.

Podophyllum hexandrum fraction (REC-2006) shows higher radioprotective efficacy in the p53-carrying hepatoma cell line: a role of cell cycle regulatory proteins.

The present study was carried out to evaluate the radioprotective efficacy of Podophyllum hexandrum fraction (REC-2006) in hepatoma cell lines having different p53 statuses. Higher radioresistance was observed in the HepG2 (p53(++)) cell line in comparison to the Hep3B (p53(-)) cell line, indicating a plausible role of p53 in radioresistance. REC-2006 exhibited nearly twice the survival in p53-expressing HepG2 cells compared with p53-negative Hep3B cells. REC-2006 treatment alone induced p53 expression as compared with untreated controls. However, REC-2006 reduced p53 expression when treated 2 hours before irradiation as compared with the irradiated HepG2 controls, indicating that REC-2006 modulates the expression of p53 to mitigate its apoptotic effect. Induction of p21 in the REC-2006 + radiation treatment group downregulated the expression of cyclin E and CDK2, leading to a delay in the G1 phase of HepG2 cells, which provided time for DNA repair or related processes. However, no significant difference in CDC2 expression in both cell lines suggested that G2 phase arrest might not be the only responsible factor for REC-2006-mediated radioprotection. Significant induction of PCNA and GADD45 expression in HepG2 cells suggested that REC-2006 increased the percentage survival of HepG2 cells by increasing the span of time as well as efficacy for repair processes. In conclusion, REC-2006 modulated the expression of p53 and thereby promoted cell cycle arrest in the G1 phase, encouraging cell proliferation and DNA repair and thus providing significantly higher protection against acute gamma-radiation in the HepG2 cell line.

Pifithrin-alpha decreases the radioprotective efficacy of a Podophyllum hexandrum Himalayan mayapple fraction REC-2006 in HepG2 cells.

Inhibition of the tumour suppressor p53 by PFT (pifithrin-alpha) promotes p53-mediated apoptosis and protects against doxorubicin-induced apoptosis. The present study was carried out to evaluate the effect of PFT on the radioprotective potential of Podophyllum hexandrum fraction (REC-2006) in HepG2 (p53++) cell line. REC-2006 (10-5 microg/ml) treatment at 2 h before irradiation (10 Gy) rendered 80+/-3% protection in HepG2 cells, whereas PFT debilitated the radioprotective potential of REC-2006. REC-2006 increased the expression of Hsp70 (heat-shock protein 70), HSF1 (heat-shock factor 1) and Bcl-2 in irradiated HepG2 cells, whereas PFT when treated with REC-2006 decreased the expression of Hsp70, HSF1 and Bcl-2 in HepG2 cells. REC-2006 facilitated post-irradiation DNA repair by pausing cell-cycle progression at G1- and G2-phase, whereas no such cell-cycle arrest was observed in irradiated HepG2 cells pretreated with PFT in irradiated HepG2 cells. No change was observed in Mdm2 (murine double minute 2) and Ras-GAP (Ras-GTPase-activating protein) expression with or without PFT treatment. Decrease in the expression of caspase 3 and Bax was observed in HepG2 cells when REC-2006 treatment was given 2 h before irradiation; however, PFT treatment increased the expression of Bax leading to apoptosis. It can be concluded that p53 expression plays a major role in the REC-2006-mediated protection against acute irradiation in HepG2 cells. PFT treatment reduced the radioprotective efficacy of REC-2006 by inhibiting the expression of HSF1 and Hsp70 and thereby the expression of Bcl-2, by up-regulating the cell-cycle-regulatory proteins and therefore reducing the span of time for DNA repair and also by inducing Bax-mediated apoptosis. PFT did not, however, show any effect on p53 regulating protein (Mdm2) and pro-survival protein (Ras-GAP).

Whole body protection to lethally irradiated mice by oral administration of semipurified fraction of Podophyllum hexandrum and post irradiation treatment of Picrorhiza kurroa.

OBJECTIVE: to evaluate the radioprotective potential of alcoholic fraction of Podophyllum hexandrum rhizomes (REC-2001) individually as well as in combination with Picrorhiza kurroa administered orally in lethally irradiated Swiss albino mice. METHODS: The study was divided into different treatment groups. Whole body survival was observed upto 30 days in all the treatment groups. Besides survival, toxicity of REC-2001 was also evaluated. All the groups were studied for spleen endogenous colony forming units (CFUs), plasma antioxidant potential and hematological variables, using standard techniques. RESULTS: Animals in radiation alone group died with in 12 days of exposure. Single dose of REC-2001 which did not bring any toxic manifestation/mortality (MTD) was found to be 155 mg/kg b.w. On administration of 250 mg/kg b.w. (single dose) 50% of the animals died (LD50), while a dose of 350 mg/kg b.w. of REC-2001 brought 100% death. Oral administration of single dose of REC-2001 (25 mg /kg b.w. -1h) prior to irradiation (10 Gy) was observed rendering up to 48% protection. Survival enhanced to the level of 55% when the animals had pre- treatment of REC-2001 (25 mg /kg b.w. -1h) followed by irradiation (10 Gy) and post treatment with a single dose of Picrorhiza kurroa rhizome extract (pkre, 8 mg/kg b.w.+1h). Radiation induced plasma antioxidant status was significantly (P < 0.02) countered by REC-2001 administration. Post treatment of pkre elevated CFU counts (P < 0.05). Total leukocytes count and hemoglobin content in REC-2001 pretreated and pkre post treated group approached normal limits within 30 days of the study. CONCLUSION: REC-2001 in combination with pkre holds promise for further studies to achieve radioprotection against lethal radiation by oral administration.

Podophyllum hexandrum (Himalayan mayapple) extract provides radioprotection by modulating the expression of proteins associated with apoptosis.

Podophyllum hexandrum Royale (Himalayan mayapple), a high-altitude Himalayan plant, has been shown to provide over 80% whole-body radioprotection in mice. To investigate the radioprotective potential of P. hexandrum at the molecular level, expression patterns of various proteins associated with apoptosis were studied in the spleen of male Swiss albino strain A mice by immunoblotting. Treatment with P. hexandrum [200 mg/kg of body weight; an ethanolic 50% (w/v) extract delivered intraperitoneally] 2 h before irradiation resulted in MAPKAP (mitogen-activated protein kinase-activated protein) kinase-2 activation along with HSF-1 (heat-shock transcription factor-1), leading to up-regulation of HSP-70 (heat-shock protein-70) as compared with sham-irradiated (10 Gy) mice. Strong inhibition of AIF (apoptosis-inducing factor) expression was observed in the mice treated with P. hexandrum 2 h before irradiation as compared with the sham-irradiated group. Inhibition in the translocation of free NF-kappaB (nuclear factor kappaB) from cytoplasm to nucleus was observed upon P. hexandrum pretreatment 2 h before irradiation when compared with radiation-treated mice. P. hexandrum pre-treatment (2 h before irradiation) resulted in inhibition of NF-kappaB translocation, and the expression of tumour suppressor protein p53 was observed to be down-regulated as compared with sham-irradiated control. An increase in the expression of proteins responsible for cell proliferation [Bcl-2 (B-cell chronic lymphocytic lymphoma 2), Ras-GAP (Ras-GTPase-activating protein) and PCNA (proliferating cell nuclear antigen)] was observed in the P. hexandrum-pretreated irradiated mice as compared with sham-irradiated controls. Caspase 3 activation resulted PARP [poly(ADP-ribose) DNA polymerase] cleavage, and DNA degradation was strongly inhibited in the mice treated with P. hexandrm (+/-irradiation) as compared with the mice treated with radiation (+/-heat shock). The present study thus clearly demonstrated that P. hexandrum extract provides protection from gamma-radiation by the modulation of expression of proteins associated with cell death.