Functional recovery after transplantation of bone marrow-derived human mesenchymal stromal cells in a rat model of spinal cord injury

Background aimsSpinal cord injury (SCI) is a medically untreatable condition for which stem cells have created hope. Pre-clinical and clinical studies have established that these cells are safe for transplantation. The dose dependency, survivability, route of administration, cell migration to injury site and effect on sensory and motor behavior in an SCI-induced paraplegic model were studied.MethodsA spinal cord contusion injury model was established in rats. Bone marrow (BM) mesenchymal stromal cells (MSC) were tagged to facilitate tracing in vivo. Two different doses (2 and 5 million cells/kg body weight) and two different routes of infusion (site of injury and lumbar puncture) were tested during and after the spinal shock period. The animals were tested post-transplantation for locomotor capacity, motor control, sensory reflex, posture and body position. Stem cell migration was observed 1 month post-transplantation in spinal cord sections.ResultsThe overall results demonstrated that transplantation of BM MSC significantly improved the locomotor and sensory behavior score in the experimental group compared with the sham control group, and these results were dose dependent. All the infused stem cells could be visualized at the site of injury and none was visualized at the injected site. This indicated that the cells had survived in vivo, were probably chemoattracted and had migrated to the lesion site.ConclusionsMSC transplanted with a lumbar puncture method migrate to the site of injury and are the most suitable for SCI healing. These cells demonstrate a dose-dependent effect and promote functional recovery when injected during or after the spinal shock period.     

Tablet Splitting of a Narrow Therapeutic Index Drug: A Case with Levothyroxine Sodium

Levothyroxine is a narrow therapeutic index, and to avoid adverse effect associated with under or excessive dosage, the dose response is carefully titrated. The tablets are marketed with a score providing an option to split. However, there are no systematic studies evaluating the effect of splitting on dose accuracy, and current study was undertaken to evaluate effects of splitting and potential causes for uniformity failures by measuring assay and content uniformity in whole and split tablets. Stability was evaluated by assaying drug for a period of 8 weeks. Effect of formulation factors on splittability was evaluated by a systematic investigation of formulation factors by preparing levothyroxine tablets in house by varying the type of excipients (binder, diluent, disintegrant, glidant) or by varying the processing factors (granulating liquid, mixing type, compression pressure). The tablets were analyzed using novel analytical tool such as near infrared chemical imaging to visualize the distribution of levothyroxine. Assay was not significantly different for whole versus split tablets irrespective of method of splitting (hand or splitter), and splitting also had no measurable impact on the stability. Split tablets either by hand or splitter showed higher rate of content uniformity failures as compared to whole tablets. Tablet splitter produced more fragmentation and, hence, more content uniformity and friability failures. Chemical imaging data revealed that the distribution of levothyroxine was heterogeneous and was dependent on type of binder and the process used in the manufacture of tablets. Splitting such tablets could prove detrimental if sub- or super-potency becomes an issue.     

Evaluation of comparative effect of pre‐ and posttreatment of selenium on mercury‐induced oxidative stress,histological alterations,and metallothionein mRNA expression in rats

To evaluate the effect of pre‐ or posttreatment of selenium (6 μmol/kg b.w., single intraperitoneal injection) in mercury intoxication, rats were exposed to mercury (12 μmol/kg b.w., single intraperitoneal injection). Exposure to mercury resulted in induced oxidative stress in liver, kidney, and brain tissues. Marked changes in serum biochemical parameters together with alterations in histopathology and an induction in metallothionein‐I and metallothionein‐II mRNA expression in the liver and kidney were observed. Pretreatment with selenium to mercury‐exposed animals had protective effect on the liver, whereas posttreatment had partial protection on restoration of altered oxidative stress parameters. In the kidney, pretreatment with selenium showed partial protection on restoration of altered biochemical parameters, whereas no protection was observed in posttreatment. The pretreatment with selenium resulted in restoration of mercury‐induced metallothionein‐I and metallothionein‐II mRNA expression, which was completely restored in the liver whereas partial restoration was observed in the kidney. Posttreatment with selenium resulted in further induction in metallothionein‐I and metallothionein‐II mRNA expression in the liver and kidney. In the brain, selenium showed partial protection on alerted biochemical parameters. Results indicate that pretreatment with selenium is beneficial in comparison to posttreatment in mercury intoxication. Thus, dietary intake of selenium within safe limit may, therefore, enable us in combating any foreseen effects due to mercury exposure. © 2010 Wiley Periodicals, Inc. J Biochem Mol Toxicol 24:123–135, 2010; Published online in Wiley InterScience ( www.interscience.wiley.com ). DOI 10.1002/jbt.20320     

Human Leukocyte Antigens and HIV Type 1 Viral Load in Early and Chronic Infection: Predominance of Evolving Relationships

Background

During untreated, chronic HIV-1 infection, plasma viral load (VL) is a relatively stable quantitative trait that has clinical and epidemiological implications. Immunogenetic research has established various human genetic factors, especially human leukocyte antigen (HLA) variants, as independent determinants of VL set-point.

Methodology/Principal Findings

To identify and clarify HLA alleles that are associated with either transient or durable immune control of HIV-1 infection, we evaluated the relationships of HLA class I and class II alleles with VL among 563 seroprevalent Zambians (SPs) who were seropositive at enrollment and 221 seroconverters (SCs) who became seropositive during quarterly follow-up visits. After statistical adjustments for non-genetic factors (sex and age), two unfavorable alleles (A*3601 and DRB1*0102) were independently associated with high VL in SPs (p<0.01) but not in SCs. In contrast, favorable HLA variants, mainly A*74, B*13, B*57 (or Cw*18), and one HLA-A and HLA-C combination (A*30+Cw*03), dominated in SCs; their independent associations with low VL were reflected in regression beta estimates that ranged from −0.47±0.23 to −0.92±0.32 log₁₀ in SCs (p<0.05). Except for Cw*18, all favorable variants had diminishing or vanishing association with VL in SPs (p≤0.86).

Conclusions/Significance

Overall, each of the three HLA class I genes had at least one allele that might contribute to effective immune control, especially during the early course of HIV-1 infection. These observations can provide a useful framework for ongoing analyses of viral mutations induced by protective immune responses.     

Cyanobacteria: potential candidates for drug discovery

Cyanobacteria are a rich source of vast array of bioactive molecules including toxins with wide pharmaceutical importance. They show varied bioactivities like antitumor, antiviral, antibacterial, antifungal, antimalarial, antimycotics, antiproliferative, cytotoxicity, immunosuppressive agents and multi-drug resistance reversers. A number of techniques are now developed and standardized for the extraction, isolation, detection and purification of cyanobacterial bioactive molecules. Some of the compounds are showing interesting results and have successfully reached to phase II and phase III of clinical trials. These compounds also serve as lead compounds for the development of synthetic analogues with improved bioactivity. Cyanobacterial bioactive molecules hold a bright and promising future in scientific research and great opportunity for drug discovery. This review mainly focuses on anticancerous, antiviral and antibacterial compounds from cyanobacteria; their clinical status; extraction and detection techniques.     

Evolutionary analysis of prokaryotic heat-shock transcription regulatory protein σ³²

Heat-stress to any living cell is known to trigger a universal defense response, called heat-shock response, with rapid induction of tens of different heat-shock proteins. Bacterial heat-shock genes are transcribed by the σ³²-bound RNA polymerase instead of the normal σ⁷⁰-bound RNA polymerase. In this study, the diversity in sequence, variation in secondary structure and function amongst the different functional regions of the proteobacterial σ³² family of proteins, and their phylogenetic relationships have been analyzed. Bacterial σ³² proteins can be subdivided into different functional regions which are referred to as regions 2, 3, and 4. There is a great deal of sequence conservation among the functional regions of proteobacterial σ³² family of proteins though some mutations are also present in these regions. Region 2 is the most conserved one, while region 4 has comparatively more variable sequences. In the present work, we tried to explore the effects of mutations in these regions. Our study suggests that the sequence diversities due to natural mutations in the different regions of proteobacterial σ³² family lead to different functions. So far, this study is the first bioinformatic approach towards the understanding of the mechanistic details of σ³² family of proteins using the protein sequence information only. This study therefore may help in elucidating the hitherto unknown molecular mechanism of the functionalities of σ³²family of proteins.     

Disintegration of Highly Soluble Immediate Release Tablets: A Surrogate for Dissolution

The purpose of the work was to investigate correlation between disintegration and dissolution for immediate release tablets containing a high solubility drug and to identify formulations where disintegration test, instead of the dissolution test, may be used as the acceptance criteria based on International Conference on Harmonization Q6A guidelines. A statistical design of experiments was used to study the effect of filler, binder, disintegrating agent, and tablet hardness on the disintegration and dissolution of verapamil hydrochloride tablets. All formulation variables, i.e., filler, binder, and disintegrating agent, were found to influence tablet dissolution and disintegration, with the filler and disintegrating agent exerting the most significant influence. Slower dissolution was observed with increasing disintegration time when either the filler or the disintegrating agent was kept constant. However, no direct corelationship was observed between the disintegration and dissolution across all formulations due to the interactions between different formulation components. Although all tablets containing sodium carboxymethyl cellulose as the disintegrating agent, disintegrated in less than 3 min, half of them failed to meet the US Pharmacopeia 30 dissolution criteria for the verapamil hydrochloride tablets highlighting the dependence of dissolution process on the formulation components other than the disintegrating agent. The results identified only one formulation as suitable for using the disintegration test, instead of the dissolution test, as drug product acceptance criteria and highlight the need for systematic studies before using the disintegration test, instead of the dissolution test as the drug acceptance criteria. The opinions expressed in this work are only of authors and do not necessarily reflect the policy and statements of the FDA.     

Residual Participation and Thermodynamic Stability Due to Molecular Interactions in IL11, IL11Rα and Gp130 from Homo sapiens: An In Silico Outlook for IL11 as a Therapeutic Remedy

International Journal of Peptide Research and Therapeutics - An extensive problem after chemotherapy is “thrombocytopenia”. It has been known (through protein assays only) to recover...     

Engineered Humicola insolens cutinase for efficient cellulose acetate deacetylation

Cutinases comprise a family of esterases with broad hydrolytic activity for chain and pendant ester groups. This work aimed to identify and improve an efficient cutinase for cellulose acetate (CA) deacetylation. The development of a mild method for CA fiber surface deacetylation will result in improved surface hydrophilicity and reactivity while, when combined with cellulases, a route to the full recycling of CA to acetate and glucose. In this study, the comparative CA deacetylation activity of four homologous wild‐type (wt) fungal cutinases from Aspergillus oryzae (AoC), Thiellavia terrestris (TtC), Fusarium solani (FsC), and Humicola insolens (HiC) was determined by analysis of CA deacetylation kinetics. wt‐HiC had the highest catalytic efficiency (≈32 [cm² L^‐1]^‐1 h^‐1). Comparison of wt‐cutinase catalytic constants revealed that differences in catalytic efficiency are primarily due to corresponding variations in corresponding substrate binding constants. Docking studies with model tetrameric substrates also revealed structural origins for differential substrate binding amongst these cutinases. Comparative docking studies of HiC point mutations led to the identification of two important rationales for engineering cutinases for CA deacetylation: (i) create a tight but not too closed binding groove, (ii) allow for hydrogen bonding in the extended region around the active site. Rationally designed HiC with amino acid substitutions I36S, predicted to hydrogen bond to CA, combined with F70A, predicted to remove steric constraints, showed a two‐fold improvement in catalytic efficiency. Continued cutinase optimization guided by a detailed understanding of structure‐activity relationships, as demonstrated here, will be an important tool to developing practical cutinases for commercial green chemistry technologies.