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1.  Duodenal perforation caused by a bird feather 
BMJ Case Reports  2013;2013:bcr2013008635.
Ingestion of gastrointestinal (GI) foreign bodies represents a challenging clinical scenario. The greater risk is at extremes of age, in those wearing dentures, alcoholics and mentally handicapped. We present a case of duodenal perforation caused by a bird feather. A 64-year-old man was presented with abdominal pain for 4 days. Abdominal examination showed signs of peritonitis. The erect abdominal x-ray showed free gas under diaphragm. Exploratory laparotomy showed purulent fluid, but no definite site of perforation could be found. So the abdomen was closed with a drain in Morison's pouch. The postoperative recovery was uneventful. He came for a repeat check-up at 4 weeks with dull aching pain in the upper abdomen and was advised for a routine upper GI endoscopy which revealed a feather penetrating the first part of the duodenum, which was removed with a foreign body removing forceps. GI foreign bodies represent a significant problem and an increased level of suspicion is important for timely diagnosis and treatment.
PMCID: PMC3603951  PMID: 23417953
2.  WHO Multidrug Therapy for Leprosy: Epidemiology of Default in Treatment in Agra District, Uttar Pradesh, India 
BioMed Research International  2015;2015:705804.
Aim. To study the magnitude of default, time of default, its causes, and final clinical outcome. Methods. Data collected in active surveys in Agra is analyzed. Patients were given treatment after medical confirmation and were followed up. The treatment default and other clinical outcomes were recorded. Results. Patients who defaulted have comparable demographic characteristics. However, among defaulters more women (62.7% in PB, 42.6% in MB) were seen than those in treatment completers (PB 52.7% and MB 35.9%). Nerve involvement was high in treatment completers: 45.7% in PB and 91.3% in MB leprosy. Overall default rate was lower (14.8%) in ROM than (28.8%) in standard MDT for PB leprosy (χ12 = 11.6, P = 0.001) and also for MB leprosy: 9.1% in ROM compared to 34.5% in MDT (χ12 = 6.0, P = 0.015). Default rate was not different (28.8% versus 34.5%, P > 0.05) in both types of leprosy given MDT. Most patients defaulted at early stage of treatment and mainly due to manageable side effects. Conclusion. The default in standard MDT both for PB and MB leprosy was observed to be significantly higher than in ROM treatment. Most defaults occurred at early stage of treatment and major contribution of default is due to side effects like drowsiness, weakness, vomiting, diarrhea, and so forth, related to poor general health. Although about half of the defaulters were observed to be cured 2.2% in PB-MDT and 10.9% of MB-MDT developed disability. This is an issue due to default. Attempts are needed to increase treatment compliance. The use of specially designed disease related health education along with easily administered drug regimens may help to reduce default.
PMCID: PMC4331159
3.  Objectively Measured Sleep and β-amyloid Burden in Older Adults: A Pilot Study 
SAGE open medicine  2014;2:10.1177/2050312114546520.
Although disturbed sleep is associated with cognitive deficits, the association between sleep disturbance and Alzheimer’s disease (AD) pathology is unclear. In this pilot study, we examined the extent to which sleep duration, sleep quality, and sleep-disordered breathing (SDB) are associated with β-amyloid (Aβ) deposition in the brains of living humans.
We studied 13 older adults (8 with normal cognition and 5 with mild cognitive impairment (MCI)). Participants completed neuropsychological testing, polysomnography and Aβ imaging with [11C]-Pittsburgh compound B.
Among participants with MCI, higher apnea-hypopnea index and oxygen desaturation index were associated with greater Aβ deposition, globally and regionally in the precuneus. There were no significant associations between SDB and Aβ deposition among cognitively normal participants. There were no significant associations between sleep duration or sleep fragmentation and Aβ deposition.
These preliminary results suggest that, among older adults with MCI, greater SDB severity is associated with greater Aβ deposition.
PMCID: PMC4304392  PMID: 25621174
sleep; sleep apnea; mild cognitive impairment; Alzheimer’s disease; amyloid; positron emission tomography
4.  Polycyclic Aromatic Hydrocarbon Residues in Serum Samples of Autopsied Individuals from Tennessee 
This study reports the concentrations of Polycyclic Aromatic Hydrocarbons (PAHs) in human blood sera samples (n = 650) obtained at autopsy from individuals who died of drug abuse, alcohol toxicity, homicide, suicide and other unknown causes. The analyzed samples from decedents revealed the presence of PAHs of which B(a)P was the most predominant one, followed by benzo(b)fluoranthene and benzo(k)fluoranthene. The other PAHs detected sporadically and measured were benzo(g,h,i)perylene, acenaphthene, anthracene, phenanthrene, and fluoranthene The mean concentrations of PAHs were greater in the twenties to fifties age groups compared to others. The PAH residue levels detected were high in African Americans compared to Caucasians, Asians, and Hispanics. It appears that environmental exposure, dietary intake and in some cases occupational exposure may have contributed to the PAH body burden. While the PAH residue concentrations measured fall within the range of those reported for healthy adults elsewhere, in isolated cases, the concentrations detected were high, calling the need for a reduction in PAH emissions and human biomonitoring studies for purposes of risk assessment.
PMCID: PMC4306864  PMID: 25547400
polycyclic aromatic hydrocarbons; benzo(a)pyrene; body burden; autopsy; serum; postmortem
5.  Molecular mechanisms involved in HIV-1 Tat-mediated induction of IL-6 and IL-8 in astrocytes 
HIV-associated neurocognitive disorders (HAND) exist in approximately 50% of infected individuals even after the introduction of highly active antiretroviral therapy. HIV-1 Tat has been implicated in HIV-associated neurotoxicity mediated through production of pro-inflammatory cytokines like IL-6 and IL-8 by astrocytes among others as well as oxidative stress. However, the underlying mechanism(s) in the up-regulation of IL-6 and IL-8 are not clearly understood. The present study was designed to determine the mechanism(s) responsible for IL-6 and IL-8 up-regulation by HIV-1 Tat.
SVG astrocytes were transiently transfected with a plasmid encoding HIV-1 Tat. The HIV-1 Tat-mediated mRNA and protein expression levels of both IL-6 and IL-8 in SVG astrocytes were quantified using real time RT-PCR and multiplex cytokine assay respectively. We also employed immunocytochemistry for staining of IL-6 and IL-8. The underlying signaling mechanism(s) were identified using pharmacological inhibitors and siRNA for different intermediate steps involved in PI3K/Akt, p38 MAPK and JNK MAPK pathways. Appropriate controls were used in the experiments and the effect of pharmacological antagonists and siRNA were observed on both mRNA expression and protein levels.
Both IL-6/IL-8 mRNA and protein showed peak expressions at 6 hours and 96 hours post-transfection, respectively. Elevated levels of IL-6/IL-8 were also confirmed by immunocytochemistry. Our studies indicated that both NF-kB and AP-1 transcription factors were involved in IL-6 and IL-8 expression mediated by HIV-1 Tat; however, AP-1 was differentially activated for either cytokine. In the case of IL-6, p38δ activated AP-1 whereas JNK but not p38 MAPK was involved in AP-1 activation for IL-8 production. On the other hand both PI3K/Akt and p38 MAPK (β subunit) were found to be involved in activation of NF-κB that led to IL-6 and IL-8 production.
Our results demonstrate HIV-1 Tat-mediated induction of both IL-6 and IL-8 in a time-dependent manner in SVG astrocytes. Furthermore, we also showed the involvement of NF-κB and AP-1 transcription factors regulated by PI3/Akt, p38 MAPK and JNK MAPK upstream signaling molecules. These results present new therapeutic targets that could be used in management of HAND.
Electronic supplementary material
The online version of this article (doi:10.1186/s12974-014-0214-3) contains supplementary material, which is available to authorized users.
PMCID: PMC4302610  PMID: 25539898
HIV-1 Tat; Astrocytes; IL-6/IL-8; NF-κB; AP-1; PI3K/Akt; p38 MAPK; JNK MAPK
6.  Identification and molecular characterization of SIV Vpr R50G mutation associated with long term survival in SIV-infected morphine dependent and control macaques 
Virology  2013;446(0):10.1016/j.virol.2013.07.027.
Viral protein R (Vpr) is an accessory protein of HIV and SIV involved in the pathogenesis of viral infection. In this study, we monitored SIV evolution in the central nervous system and other organs from morphine-dependent and control animals by sequencing vpr in an attempt to understand the relationship between drug abuse, disease progression, and compartmentalization of viral evolution. Animals in the morphine group developed accelerated disease and died within twenty weeks post-infection. A unique mutation, R50G, was identified in the macaques that survived regardless of morphine exposure. Functional studies revealed that the R50G mutation exhibited altered cellular localization and decreased the expression levels of both IL-6 and IL-8. Our results, therefore, suggest that sequence changes within the SIV/17E-Fr vpr occur regardless of drug abuse but correlate with survival, and that they alter disease progression rates by affecting Vpr functions.
PMCID: PMC3844929  PMID: 24074576
HIV; SIV; vpr; viral evolution; morphine; brain; PBMC; proviral; IL-6; IL-8
7.  Methamphetamine Alters the Normal Progression by Inducing Cell Cycle Arrest in Astrocytes 
PLoS ONE  2014;9(10):e109603.
Methamphetamine (MA) is a potent psychostimulant with a high addictive capacity, which induces many deleterious effects on the brain. Chronic MA abuse leads to cognitive dysfunction and motor impairment. MA affects many cells in the brain, but the effects on astrocytes of repeated MA exposure is not well understood. In this report, we used Gene chip array to analyze the changes in the gene expression profile of primary human astrocytes treated with MA for 3 days. Range of genes were found to be differentially regulated, with a large number of genes significantly downregulated, including NEK2, TTK, TOP2A, and CCNE2. Gene ontology and pathway analysis showed a highly significant clustering of genes involved in cell cycle progression and DNA replication. Further pathway analysis showed that the genes downregulated by multiple MA treatment were critical for G2/M phase progression and G1/S transition. Cell cycle analysis of SVG astrocytes showed a significant reduction in the percentage of cell in the G2/M phase with a concomitant increase in G1 percentage. This was consistent with the gene array and validation data, which showed that repeated MA treatment downregulated the genes associated with cell cycle regulation. This is a novel finding, which explains the effect of MA treatment on astrocytes and has clear implication in neuroinflammation among the drug abusers.
PMCID: PMC4188627  PMID: 25290377
8.  π vs σ-Radical States of One-Electron Oxidized DNA/RNA Bases: A Density Functional Theory Study 
The journal of physical chemistry. B  2013;117(39):11623-11632.
As a result of their inherent planarity, DNA base radicals generated by one electron oxidation/reduction or bond cleavage form π- or σ-radicals. While most DNA base systems form π-radicals there are a number of nucleobase analogs such as one-electron oxidized 6-azauraci1, 6-azacytosine, and 2-thiothymine or one-electron reduced 5-bromouracil that form more reactive σ-radicals. Elucidating the availability of these states within DNA, base radical electronic structure is important to the understanding of the reactivity of DNA base radicals in different environments. In this work, we address this question by the calculation of the relative energies of π- and σ-radical states in DNA/RNA bases and their analogs. We used density functional theory B3LYP/6-31++G** method to optimize the geometries of π- and σ-radicals in Cs symmetry (i.e., planar) in the gas phase and in solution using the polarized continuum model (PCM). The calculations predict that σ- and π-radical states in one electron oxidized bases of thymine, T(N3-H)•, and uracil, U(N3-H)• are very close in energy, i.e., the π-radical is only ca. 4 kcal/mol more stable than the σ-radical. For the one electron oxidized radicals of cytosine, C•+, C(N4-H)•, adenine, A•+, A(N6-H)•, and guanine, G•+, G(N2-H)•, G(N1-H)• the π-radicals are ca. 16 to 41 kcal/mol more stable than their corresponding σ-radicals. Inclusion of solvent (PCM) is found to stabilize the π- over σ-radical of each of the systems. U(N3-H)• with three discrete water molecules in the gas phase, is found to form a three-electron σ bond between N3 atom of uracil and O atom of a water molecule but on inclusion of full solvation and discrete hydration the π-radical remains most stable..
PMCID: PMC3946539  PMID: 24000793
DNA base radical; sigma-radical; pi-radical; radical of DNA bases; DFT; cation radical of bases
9.  Synthesis and Antiproliferative Activities of Quebecol and Its Analogs 
Bioorganic & medicinal chemistry letters  2013;23(19):10.1016/j.bmcl.2013.07.058.
Simple and efficient synthesis of quebecol and a number of its analogs was accomplished in five steps. The synthesized compounds were evaluated for antiproliferative activities against human cervix adenocarcinoma (HeLa), human ovarian carcinoma (SKOV-3), human colon carcinoma (HT-29), and human breast adenocarcinoma (MCF-7) cancer cell lines. Among all the compounds, 7c, 7d, 7f, and 8f exhibited antiproliferative activities against four tested cell lines with inhibition over 80% at 75 μM after 72 h, whereas, compound 7b and 7g were more selective towards MCF-7 cell line. The IC50 values for compounds 7c, 7d, and 7f were 85.1 μM, 78.7 μM, and 80.6 μM against MCF-7 cell line, respectively, showing slightly higher antiproliferactive activtiy than the synthesized and isolated quebecol with an IC50 value of 104.2 μM against MCF-7.
PMCID: PMC3816661  PMID: 23953195
10.  Recurrent primary retro-bulbar hydatid cysts 
Retro-bulbar hydatid cysts are extremely uncommon, while nonorbital forms are frequently encountered disease in underdeveloped countries. Most of these are situated in the superolateral and superomedial angle of the orbit. We report a case of recurrent primary hydatid cysts of the orbit, situated in different locations in the orbit. A 35-year-old female patient was admitted to Department of Neurosurgery with proptosis, ptosis and watering from left eye. She also complained for headache with excruciating pain in left eye and loss of vision in left eye. Neurological examination revealed limited ocular mobility in all directions. Visual acuity was reduced to finger counting at 2-feet distance. Papilledema was found in ophthalmic examination. This case was considered as recurrence of primary infection because there was no previous history of hydatid disease and no finding of liver and lung cysts on radiological examinations. Treatment of orbital hydatid cyst, early diagnosis, surgical excision and systemic use of albendazole are suggested.
PMCID: PMC4323980
Ocular mobility; papilledema; proptosis; ptosis; retro-bulbar hydatid cyst; visual acuity
11.  Copper triflate-mediated synthesis of 1,3,5-triarylpyrazoles in [bmim][PF6] ionic liquid and evaluation of their anticancer activities 
RSC advances  2013;3(35):15396-15403.
A simple, efficient, and environment friendly protocol for the synthesis of 1,3,5-triarylpyrazole and 1,3,5-triarylpyrazolines in [bimm][PF6] ionic liquid mediated by Cu(OTf)2 is described. The reaction protocol gave 1,3,5-triarylpyrazoles in good to high yields (71-84%) via a one-pot addition–cyclocondensation between chalcones and arylhydrazines, and oxidative aromatization without requirement for an additional oxidizing reagent. The catalyst can be reused up to four cycles without much loss in the catalytic activity. The pyrazoles (4a-o) and pyrazolines (3a-n) were evaluated for antiproliferative activity in SK-OV-3, HT-29, and HeLa human cancer cells lines. Among all compounds, 3b inhibited cell proliferation of HeLa cells by 80% at a concentration of 50 μM.
PMCID: PMC3806107  PMID: 24163734
12.  Formation of S-Cl phosphorothioate adduct radicals in dsDNA-S-oligomers: Hole transfer to guanine vs. disulfide anion radical formation 
Journal of the American Chemical Society  2013;135(34):12827-12838.
In phosphorothioate containing dsDNA-oligomers (S-oligomers), one of the two non-bridging oxygen atoms in the phosphate moiety of sugar-phosphate backbone is replaced by sulphur. In this work, electron spin resonance (ESR) studies of one-electron oxidation of several S-oligos by Cl2•− at low temperatures are investigated. Electrophilic addition of Cl2•− to phosphorothioate with elimination of Cl− leads to the formation of a 2-center three-electron σ2σ*1 bonded adduct radical (-P-S∸Cl). In AT S-oligomers with mutiple phosphorothioates, i.e., d[ATATAsTsAsT]2, -P-S∸Cl reacts with a neighboring phosphorothioate to form the σ2σ*1 bonded disulphide anion radical ([-P-S∸S-P-]−). With AT S-oligomers with a single phosphorothioate, i.e., d[ATTTAsAAT]2, reduced levels of conversion of -P-S∸Cl dsDNA [-P-S∸S-P-]− are found. For guanine containing S-oligomers containing one phosphorothioate, -P-S∸Cl results in one-electron oxidation of guanine base but not of A, C, or T thereby leading to selective hole transfer to G. The redox potential of -P-S∸Cl is thus higher than that of G but is lower than those of A, C, and T. Spectral assignments to -P-S∸Cl and [-P-S∸S-P-]− are based on reaction of Cl2•− with the model compound diisopropyl phosphorothioate. The results found for d[TGCGsCsGCGCA]2 suggest that [-P-S∸S-P-]− undergoes electron transfer to the one-electron oxidized G healing the base but producing a cyclic disulfide bonded backbone with a substantial bond strength (50 kcal/mol). Formation of -P-S∸Cl and its conversion to [-P-S∸S-P-]− is found to be unaffected by O2 and this is supported by the theoretically calculated electron affinities and reduction potentials of [-P-S-S-P-] and O2.
PMCID: PMC3789529  PMID: 23885974
13.  Transcriptome Wide Identification and Validation of Calcium Sensor Gene Family in the Developing Spikes of Finger Millet Genotypes for Elucidating Its Role in Grain Calcium Accumulation 
PLoS ONE  2014;9(8):e103963.
In finger millet, calcium is one of the important and abundant mineral elements. The molecular mechanisms involved in calcium accumulation in plants remains poorly understood. Transcriptome sequencing of genetically diverse genotypes of finger millet differing in grain calcium content will help in understanding the trait.
Principal Finding
In this study, the transcriptome sequencing of spike tissues of two genotypes of finger millet differing in their grain calcium content, were performed for the first time. Out of 109,218 contigs, 78 contigs in case of GP-1 (Low Ca genotype) and out of 120,130 contigs 76 contigs in case of GP-45 (High Ca genotype), were identified as calcium sensor genes. Through in silico analysis all 82 unique calcium sensor genes were classified into eight calcium sensor gene family viz., CaM & CaMLs, CBLs, CIPKs, CRKs, PEPRKs, CDPKs, CaMKs and CCaMK. Out of 82 genes, 12 were found diverse from the rice orthologs. The differential expression analysis on the basis of FPKM value resulted in 24 genes highly expressed in GP-45 and 11 genes highly expressed in GP-1. Ten of the 35 differentially expressed genes could be assigned to three documented pathways involved mainly in stress responses. Furthermore, validation of selected calcium sensor responder genes was also performed by qPCR, in developing spikes of both genotypes grown on different concentration of exogenous calcium.
Through de novo transcriptome data assembly and analysis, we reported the comprehensive identification and functional characterization of calcium sensor gene family. The calcium sensor gene family identified and characterized in this study will facilitate in understanding the molecular basis of calcium accumulation and development of calcium biofortified crops. Moreover, this study also supported that identification and characterization of gene family through Illumina paired-end sequencing is a potential tool for generating the genomic information of gene family in non-model species.
PMCID: PMC4144799  PMID: 25157851
14.  VMAT2 and dopamine neuron loss in a primate model of Parkinson’s disease 
Journal of neurochemistry  2007;105(1):78-90.
We used positron emission tomography (PET) to measure the earliest change in dopaminergic synapses and glial cell markers in a chronic, low-dose MPTP non-human primate model of Parkinson’s disease (PD). In vivo levels of dopamine transporters (DAT), vesicular monoamine transporter-type 2 (VMAT2), amphetamine-induced dopamine release (AMPH-DAR), D2-dopamine receptors (D2R) and translocator protein 18 kDa (TSPO) were measured longitudinally in the striatum of MPTP-treated animals. We report an early (2 months) decrease (46%) of striatal VMAT2 in asymptomatic MPTP animals that preceded changes in DAT, D2R, and AMPH-DAR and was associated with increased TSPO levels indicative of a glial response. Subsequent PET studies showed progressive loss of all pre-synaptic dopamine markers in the striatum with expression of parkinsonism. However, glial cell activation did not track disease progression. These findings indicate that decreased VMAT2 is a key pathogenic event that precedes nigrostriatal dopamine neuron degeneration. The loss of VMAT2 may result from an association with α-synuclein aggregation induced by oxidative stress. Disruption of dopamine sequestration by reducing VMAT2 is an early pathogenic event in the dopamine neuron degeneration that occurs in the MPTP non-human primate model of PD. Genetic or environmental factors that decrease VMAT2 function may be important determinants of PD.
PMCID: PMC4137401  PMID: 17988241
MPTP; non-human primate; Parkinson’s disease; PET; translocator protein 18 kDa; VMAT2
15.  Excited state Proton-coupled electron transfer in 8-oxoG-C and 8-oxoG-A base pairs: A time dependent density functional theory (TD-DFT) study 
In a recent experiment, the repair efficiency of DNA thymine cyclobutane dimers (T<>T) on UV excitation of 8-oxoG base paired either to C or A was reported. An electron transfer mechanism from an excited charge transfer state of 8-oxoG-C (or 8-oxoG-A) to T<>T was proposed and 8-oxoG-A was found to be 2 – 3 times more efficient than 8-oxoG-C in repair of T<>T. Intra base pair proton transfer (PT) in charge transfer (CT) excited states of the base pairs was proposed to quench the excited state and prevent T<>T repair. In this work, we investigate this process with TD-DFT calculations of the excited states of 8-oxoG-C and 8-oxoG-A base pairs in the Watson-Crick and Hoogsteen base pairs using long-range corrected density functional, ωB97XD/6–31G* method. Our gas phase calculations showed that CT excited state (1ππ*(CT)) of 8-oxoG-C appears at lower energy than the 8-oxoG-A. For 8-oxoG-C, TD-DFT calculations show the presence of a conical intersection (CI) between the lowest 1ππ*(PT-CT) excited state and the ground state which likely deactivates the CT excited state via a proton-coupled electron transfer (PCET) mechanism. The 1ππ*(PT-CT) excited state of 8-oxoG-A base pair lies at higher energy and its crossing with ground state is inhibited because of a high energy gap between 1ππ*(PT-CT) excited state and ground state. Thus the gas phase calculations suggest the 8-oxoG-A would have longer excited state lifetimes. When the effect of solvation is included using the PCM model, both 8-oxoG-A and 8-oxoG-C show large energy gaps between the ground state and both the excited CT and PT-CT states and suggest little difference would be found between the two base pairs in repair of the T<>T lesion. However, in the FC region the solvent effect is greatly diminished owing to the slow dielectric response time and smaller gaps would be expected.
PMCID: PMC3717295  PMID: 23478652
8-oxo-7,8-dihydroguanine (8-oxoG); excited state; proton-coupled electron transfer; PCET; excited state electron-induced proton transfer; density functional; thymine dimer (T<>T)
16.  Methamphetamine toxicity and its implications during HIV-1 infection 
Journal of neurovirology  2011;17(5):401-415.
Over the past two decades methamphetamine (MA) abuse has seen a dramatic increase. The abuse of MA is particularly high in groups that are at higher risk for HIV-1 infection, especially men who have sex with men (MSM). This review is focused on MA toxicity in the CNS as well as in the periphery. In the CNS, MA toxicity is comprised of numerous effects, including, but not limited to, oxidative stress produced by dysregulation of the dopaminergic system, hyperthermia, apoptosis, and neuroinflammation. Multiple lines of evidence demonstrate that these effects exacerbate the neurodegenerative damage caused by CNS infection of HIV perhaps because both MA and HIV target the frontostriatal regions of the brain. MA has also been demonstrated to increase viral load in the CNS of SIV-infected macaques. Using transgenic animal models, as well as cultured cells, the HIV proteins Tat and gp120 have been demonstrated to have neurotoxic properties that are aggravated by MA. In addition, MA has been shown to exhibit detrimental effects on the blood–brain barrier (BBB) that have the potential to increase the probability of CNS infection by HIV. Although the effects of MA in the periphery have not been as extensively studied as have the effects on the CNS, recent reports demonstrate the potential effects of MA on HIV infection in the periphery including increased expression of HIV co-receptors and increased expression of inflammatory cytokines.
PMCID: PMC4118146  PMID: 21786077
Methamphetamine toxicity; HIV-1 infection; MSM; CNS
17.  A LC-MS/MS Method for Concurrent Determination of Nicotine Metabolites and Role of CYP2A6 in Nicotine Metabolism in U937 Macrophages: Implications in Oxidative Stress in HIV + Smokers 
Nicotine, the major constituent of tobacco, is predominantly metabolized by liver CYP2A6 into cotinine and many other compounds, including nicotine-derived nitrosamine ketone (NNK), which is known to cause oxidative stress. We have recently shown that CYP2A6 is highly expressed in U937 monocyte-derived macrophages. In this study we investigated the role of CYP2A6 in nicotine metabolism and oxidative stress in U937 macrophages. To study nicotine metabolism, we developed a highly sensitive LC-MS/MS method for simultaneous quantitative determination of nicotine, cotinine, and NNK. The LC-MS/MS analysis was carried out by multiple reaction monitoring mass transitions with m/z of 163.2/130.1, 177.4/98.3, and 208.4/122.1 for nicotine, cotinine, and NNK, respectively. The calibration curves were linear within 3.3–1028.1 ng/ml for nicotine and 0.3–652.6 ng/ml for cotinine and NNK. This novel method was then applied to quantify nicotine metabolites, cotinine and NNK, in nicotine-treated U937 macrophages. Cotinine and NNK initially formed at 30 min, followed by a peak at 2–3 h. The role of CYP2A6 in nicotine metabolism in U937 macrophages was further confirmed by using CYP2A6-selective inhibitor, tryptamine, which significantly decreased cotinine (70%) and completely inhibited NNK formations. Finally, we showed that nicotine-treated macrophages increase the formation of oxidant at 30–60 min, which is consistent with the initial formation of cotinine and NNK. In conclusion, we have developed a new LCMS/MS method for concurrent determination of nicotine metabolites and analyzed the role of CYP2A6 in nicotine metabolism and oxidative stress in U937 macrophages, which may have implications in viral replication among HIV+smokers.
PMCID: PMC4118724  PMID: 21655912
Nicotine; CYP2A6; Oxidative stress; Macrophages; HIV-1; LC-MS/MS
18.  Morphine and Rapid Disease Progression in Nonhuman Primate Model of AIDS: Inverse Correlation Between Disease Progression and Virus Evolution 
HIV and simian immunodeficiency virus (SIV) have a formidable capacity for mutation and adaptation, a characteristic that has contributed to the extensive genetic variability. Evolutionary pressures imposed within the host and the viral capacity to mutate lead to the generation of such variants. To date, very little information is available regarding the evolution of HIV with drug abuse as a cofounding factor. Using our macaque model of drug dependency and AIDS, we have investigated the dynamics of SIV mutations in the genes tat, vpr, envelope, and nef. The results presented in this review, from our laboratory and others, contribute to the overall understanding of how drugs of abuse might influence immune selective pressure contribution to variation in different SIV genes. Additionally, the studies presented could help enlighten the development of HIV vaccines that take into consideration viral diversity.
PMCID: PMC4095870  PMID: 20013315
morphine; SIV; viral evolution; AIDS
19.  New approaches to the inhibition of replication of viral pathogens 
This meeting was a special symposium sponsored by the American Society for Biochemistry and Molecular Biology. The conference was held in Gangzhou, China on July 24–26, 2011 and shared a venue with the Society of Chinese Bioscientists in America (SCBA) Thirteenth International Symposium. Over 150 participants from the Americas, Europe, Asia and Australia attended the meeting. The meeting report focuses on two areas of research in which there have been exciting developments that have application to the development of antivirals: the regulation of host and viral mRNA by RNAi and NF-kB regulation of viral gene expression.
PMCID: PMC4090758  PMID: 22029515
20.  HIV-1 and Alcohol: Interactions in the Central Nervous System 
The use of alcohol has been associated with both an increased risk of acquisition of HIV-1 infection and an increased rate of disease progression among those already infected by the virus. The potential for alcohol to exacerbate the effects of HIV infection is especially important in the CNS because this area is vulnerable to the combined effects of alcohol and HIV infection. The effects of alcohol on glial cells are mediated through receptors such as TLR4 and NMDAR. This causes the activation of signaling molecules such as IRAK and various members of the p38MAPK family and subsequent activation of transcription factors such as NF-κB and AP-1. The eventual outcome is an increase in pro-inflammatory cytokine production by glial cells. Alcohol also induces higher levels of NADPH oxidase in glial cells which leads to an increased production of ROS. Viral invasion of the CNS occurs early after infection, and HIV proteins have also been demonstrated to increase levels of pro-inflammatory cytokines and ROS in glial cells through activation of some of the same pathways activated by alcohol. Both cell culture systems and animal models have demonstrated that concomitant exposure to alcohol and HIV/HIV proteins results in increased levels of expression of pro-inflammatory cytokines such as IL-1β, and TNF-α, along with increased levels of oxidative stress. Clinical studies also suggest that alcohol exacerbates the CNS effects of HIV-1 infection. This review focuses on the mechanisms by which alcohol causes increased CNS damage in HIV-1-infection.
PMCID: PMC4086304  PMID: 24134164
Alcohol; HIV-1; Central Nervous System
21.  HIV-1 gp120 and Drugs of Abuse: Interactions in the Central Nervous System 
Current HIV research  2012;10(5):369-383.
HIV-1 infection is a global public health problem with more than 34 million people living with HIV infection. Although great strides have been made in treating this epidemic with therapeutic agents, the increase in patient life span has been coincident with an increase in the prevalence of HIV-associated neurocognitive disorders (HAND). HAND is thought to result from the neurotoxic effects of viral proteins that are shed from HIV-infected microglial cells. One of the primary neurotoxins responsible for this effect is the HIV-1 glycoprotein gp120. Exposure of neurons to gp120 has been demonstrated to cause apoptosis in neurons, as well as numerous indirect effects such as an increase in inflammatory cytokines, an increase in oxidative stress, and an increase in permeability of the blood-brain barrier. In many patients, the use of drugs of abuse (DOA) exacerbates the neurotoxic effects of gp120. Cocaine, methamphetamine and morphine are three DOAs that are commonly used by those infected with HIV-1. All three of these DOAs have been demonstrated to increase oxidative stress in the CNS as well as to increase permeability of the blood-brain barrier. Numerous model systems have demonstrated that these DOAs have the capability of exacerbating the neurotoxic effects of gp120. This review will summarize the neurotoxic effects of gp120, the deleterious effects of cocaine, methamphetamine and morphine on the CNS, and the combined effects of gp120 in the context of these drugs.
PMCID: PMC4086306  PMID: 22591361
CNS; cocaine; gp120; HIV; methamphetamine; morphine; central nervous system; HAND; drug of abuse; ARV
22.  Opiates, immune system, acquired immunodeficiency syndrome, and nonhuman primate model 
Journal of neurovirology  2008;14(4):279-285.
Both human immunodeficiency virus (HIV) and illicit drug addiction remain major health problems not only in the United States but all over globe. The effect of drug addiction on HIV/AIDS (acquired immunodeficiency syndrome) has been somewhat underexplored. However, in United States more than one fourth of HIV-positive individuals are injection drug users. Opiates are known to negatively affect the immune system, and therefore may have deleterious effects on progression of disease among HIV-infected individuals. This review discusses the effects of opiates on immune system as well as its effect on HIV replication and AIDS progression. In addition, the effects of opiates on disease progression in non-human primate model of AIDS is presented with at least one possible reason for rapid disease progression in multi-virus the challenge model.
PMCID: PMC4086310  PMID: 18780228
HIV; morphine; macaque; SIV; SHIV
23.  Differential Effects of Ethanol on Spectral Binding and Inhibition of Cytochrome P450 3A4 with Eight Protease Inhibitors Antiretroviral Drugs 
Cytochrome P450 3A4 (CYP3A4) is the most abundant CYP enzyme in the liver, which metabolizes approximately 50% of the marketed drugs including antiretroviral agents. CYP3A4 induction by ethanol and its impact on drug metabolism and toxicity is known. However, CYP3A4–ethanol physical interaction and its impact on drug binding, inhibition, or metabolism is not known, except that we have recently shown that ethanol facilitates the binding of a protease inhibitor (PI), nelfinavir, with CYP3A4. The current study was designed to examine the effect of ethanol on spectral binding and inhibition of CYP3A4 with all currently used PIs that differ in physicochemical properties.
We performed type I and type II spectral binding with CYP3A4 at 0 and 20 mM ethanol and varying PIs’ concentrations. We also performed CYP3A4 inhibition using 7-benzyloxy-4-trifluoromethylcoumarin substrate and NADPH at varying concentrations of PIs and ethanol.
Atazanavir, lopinavir, saquinavir, and tipranavir showed type I spectral binding, whereas indinavir and ritonavir showed type II. However, amprenavir and darunavir did not show spectral binding with CYP3A4. Ethanol at 20 mM decreased the maximum spectral change (δAmax) with type I lopinavir and saquinavir, but it did not alter δAmax with other PIs. Ethanol did not alter spectral binding affinity (KD) and inhibition constant (IC50) of type I PIs. However, ethanol significantly decreased the IC50 of type II PIs, indinavir and ritonavir, and markedly increased the IC50 of amprenavir and darunavir.
Overall, our results suggest that ethanol differentially alters the binding and inhibition of CYP3A4 with the PIs that have different physicochemical properties. This study has clinical relevance because alcohol has been shown to alter the response to antiretroviral drugs, including PIs, in HIV-1-infected individuals.
PMCID: PMC4082664  PMID: 21682753
CYP3A4; Alcohol; Protease Inhibitors; Spectral Binding; Inhibition
24.  Early rehabilitation after surgery program versus conventional care during perioperative period in patients undergoing laparoscopic assisted total gastrectomy 
Journal of Minimal Access Surgery  2014;10(3):132-138.
To evaluate the safety and efficacy of early rehabilitation after surgery program (ERAS) in patients undergoing laparoscopic assisted total gastrectomy.
This is a study where 47 patients who are undergoing lap assisted total gastrectomy are selected. Twenty-two (n = 22) patients received enhanced recovery programme (ERAS) management and rest twenty-five (n = 25) conventional management during the perioperative period. The length of postoperative hospital stay, time to passage of first flatus, intraoperative and postoperative complications, readmission rate and 30 day mortality is compared. Serum levels of C-reactive protein pre-operatively and also on post-op day 1 and 3 are compared.
Postoperative hospital stay is shorter in ERAS group (78 ± 26 h) when compared to conventional group (140 ± 28 h). ERAS group passed flatus earlier than conventional group (37 ± 9 h vs. 74 ± 16 h). There is no significant difference in complications between the two groups. Serum levels of CRP are significantly low in ERAS group in comparison to conventional group. [d1 (52.40 ± 10.43) g/L vs. (73.07 ± 19.32) g/L, d3 (126.10 ± 18.62) g/L vs. (160.72 ± 26.18) g/L)].
ERAS in lap-assisted total gastrectomy is safe, feasible and efficient and it can ameliorate post-operative stress and accelerate postoperative rehabilitation in patients with gastric cancer. Short term follow up results are encouraging but we need long term studies to know its long term benefits.
PMCID: PMC4083545  PMID: 25013329
Early rehabilitation after surgery (ERAS); fast track surgery; gastric cancer; perioperative period; total gastrectomy
25.  Randomised study on single stage laparo-endoscopic rendezvous (intra-operative ERCP) procedure versus two stage approach (Pre-operative ERCP followed by laparoscopic cholecystectomy) for the management of cholelithiasis with choledocholithiasis 
Journal of Minimal Access Surgery  2014;10(3):139-143.
The ‘Rendezvous’ technique consists of laparoscopic cholecystectomy (LC) standards with intra-operative cholangiography followed by endoscopic sphincterotomy. The sphincterotome is driven across the papilla through a guidewire inserted by the transcystic route. In this study, we intended to compare the two methods in a prospective randomised trial.
From 2005 to 2012, we enrolled 83 patients with a diagnosis of cholecysto-choledocolithiasis. They were randomised into two groups. In ‘group-A’,41 patients were treated with two stages management, first by pre-operative endoscopic retrograde cholangiopancreatography (ERCP) and common bile duct (CBD) clearance and second by LC. In ‘group-B’, 42 patients were treated with LC and intra-operative cholangiography; and when diagnosis of choledocholithiasis was confirmed, patients had undergone one stage management of by Laparo-endoscopic Rendezvous technique.
In arm-A and arm-B groups, complete CBD clearance was achieved in 29 and 38 patients, respectively. Failure of the treatment in arm-A was 29% and in arm-B was 9.5%. In arm-A, selective CBD cannulation was achieved in 33 cases (80.5%) and in arm-B in 39 cases (93%). In arm-Agroup, post-ERCP hyperamylasia was presented in nine patients (22%) and severe pancreatitis in five patients (12%) versus none of the patients (0%) in arm-B group, respectively. Mean post-operative hospital stay in arm-A and arm-B groups are 10.9 and 6.8 days, respectively.
One stage laparo-endoscopic rendezvous approach increases selective cannulation of CBD, reduces post-ERCP pancreatitis, reduces days of hospital stay, increases patient's compliance and prevents unnecessary intervention to CBD.
PMCID: PMC4083546  PMID: 25013330
Endoscopic retrograde cholangiopancreaticography; laparoscopic cholecystectomy; rendezvous technique; sphincterotomy

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