Comparative study of bone marrow and blood plasma levels of IL-2 in aplastic anaemia and their relationship with disease severity.

OBJECTIVES: Interleukin-2 (alias: IL-2, TCGF, Lymphokine), a type of interleukin, is also a potent signalling molecule in the signalling cascade of the immune-mediated activation of T Lymphocytes leading to the destruction of haematopoietic stem cell (HSC) which is the basis of acquired aplastic anaemia (AAA). The objective was to study the association of IL-2 in the bone marrow plasma (BMP) and peripheral blood plasma (PBP) in AAA patients. METHODS: A total of 52 BMP and PBP-paired samples (both from the same patients) was collected from the confirmed AAA patients and 10 healthy individuals. The level of IL-2 was measured by the quantitative enzyme-linked immunosorbent assay (ELISA). The Mann-Whitney U test was used for statistical analysis. RESULTS: Significantly increased level of IL-2 was observed in the BMP than PBP of AAA patients. The level of IL-2 in PBP and BMP was found to be very low in the control cases. Considerably increased levels of IL-2 were found in the PBP and BMP of AAA patients as compared to controls (48.54 ± 21.89 vs. 1.99 ± 1.25 p-value < 0.00001) and (75.33 ± 41.9 vs. 3.12 ± 1.82; p-value < 0.00001) respectively. Among these patients, the IL-2 levels were higher in patients with Very Severe Aplastic Anaemia (VSAA) and Severe Aplastic Anaemia (SAA) than those with Non-severe Aplastic Anaemia (NSAA) in the PBP (65.6 ± 23.61 vs. 31.72 ± 7.64; p-value 0.00338) and (45.37 ± 16.25 vs. 31.72 ± 7.64; p-value 0.01468) respectively. Again the IL-2 levels were higher in patients with VSAA and SAA than those with NSAA in the BMP (115.01 ± 38.91 vs. 38.32 ± 19.49; p-value < 0.00001) and (66.44 ± 23.34 vs. 38.32 ± 19.49; p-value 0.0006). The IL-2 level was higher in VSAA than SAA in PBP (65.6 ± 23.61vs. 45.37 ± 16.25; p-value 0.0114) and BMP (115.01 ± 38.91 vs. 66.44 ± 23.34; p-value 0.00044). CONCLUSION: This study emphasized on the bone marrow and blood plasma levels of IL-2 in aplastic anaemia and their relationship with disease severity. The results indicate towards the fact that IL-2 may have an important association with the marrow failure of AAA patients and thus can help in disease development. Further study is necessary for better understanding.

Amelioration of Pain on Injection of Propofol: A Comparison of Pretreatment with Granisetron Vs Lignocaine.

INTRODUCTION: Pain during propofol injection is a very commonly and frequently encountered event during induction of anaesthesia. A 5HT3 antagonists like granisetron are commonly used just prior to intravenous propofol as pre anaesthetic medication to prevent emesis in patients. AIM: Comparison of pre treatment with granisetron versus lignocaine with respect to amelioration of pain induced by injection of propofol, in patient admitted for elective surgery with general anaesthesia. MATERIALS AND METHODS: A randomized double blinded controlled study was conducted with patients divided into three groups with (n=30) in each group. Group I (the placebo group) received 2 ml of 0.9% normal saline, Group II received 2 ml of 1% lignocaine and Group III received 2 ml of granisetron (1 mg/ml) as pre treatment medication respectively. The patient's complain regarding pain on intravenous propofol administration was recorded using the Verbal Rating Score. Pulse, BP, SpO2 were noted meticulously on three occasions-immediately after pre-treatment, injecting full dose of propofol (not for pain assessment) and after 10 minutes. The results were analysed using the null hypothesis and two sample t-tests. RESULTS: It was observed and obvious that the relief of pain was significant (p<0.05) when granisetron or lignocaine was compared with the placebo group. But there was insignificant difference (p>0.05) when granisetron was compared with lignocaine in terms of relieve of pain induced by propofol. CONCLUSION: It was concluded that parenteral administration of granisetron can be considered to be superior to lignocaine as pre treatment medication for pain relief after propofol injection along with the advantage of its anti-emetic effect.

The cytoplasmic domain is critical to the tumor suppressor activity of TSLC1 in non-small cell lung cancer.

The tumor suppressor gene in lung cancer (TSLC1) encodes a membrane glycoprotein containing extensive homology in the extracellular domain with the immunoglobulin-superfamily cell adhesion molecules. The intracellular cytoplasmic domain (CT) contains a protein 4.1 (FERM) binding motif, and a PDZ-interacting motif. Expression of TSLC1 is silenced in non-small cell lung cancer and in other cancers by promoter hypermethylation. Restoration of TSLC1 expression suppresses tumorigenicity of lung cancer cells. We report here the critical role of the FERM-binding and PDZ- interacting domains of TSLC1 in tumor suppressor activity in non-small cell lung cancer. The entire CT domain [amino acid (aa) 398-442], the FERM binding motif (aa 398-410), or the PDZ-interacting motif (aa 432-442) was deleted to generate mutants CT1, CT3, and CT4, respectively. The lung cancer cell line A549, deficient in TSLC1 expression, was stably transfected with the wild-type TSLC1 or the deletion mutants. The cell lines were then injected into athymic (nu/nu) nude mice, and tumor formation at the sites of injection was monitored. A549 cells stably transfected with the empty vector or mutant TSLC1 constructs induced tumors at the sites of injection within 10 days. In contrast, A549 cells expressing wild-type TSLC1 showed the appearance of tumors after 35 days, and the tumors grew substantially slower. A549 cells expressing wild-type TSLC1 also showed suppression of anchorage-independent colony formation in soft agar and markedly increased cell-cell adhesion activity. These results suggest that the cytoplasmic domain of TSLC1 is important in its tumor suppressor activity, and the tumor suppression activity involve protein(s) interacting with the FERM- and PDZ-interacting regions.

The membrane-proximal region of vesicular stomatitis virus glycoprotein G ectodomain is critical for fusion and virus infectivity.

The glycoprotein (G) of vesicular stomatitis virus (VSV) is responsible for binding of virus to cells and for mediating virus entry following endocytosis by inducing fusion of the viral envelope with the endosomal membrane. The fusion peptide of G is internal (residues 116 to 137) and exhibits characteristics similar to those of other internal fusion peptides, but recent studies have implicated the region adjacent to the transmembrane domain as also being important for G-mediated membrane fusion. Sequence alignment of the membrane-proximal region of G from several different vesiculoviruses revealed that this domain is highly conserved, suggesting that it is important for G function. Mutational analysis was used to show that this region is not essential for G protein oligomerization, transport to the cell surface, or incorporation into virus particles but that it is essential for acid-induced membrane fusion activity and for virus infectivity. Deletion of the 13 membrane-proximal amino acids (N449 to W461) dramatically reduced cell-cell fusion activity and reduced virus infectivity approximately 100-fold, but mutation of conserved aromatic residues (W457, F458, and W461) either singly or together had only modest effects on cell-cell fusion activity; recombinant virus encoding these mutants replicated as efficiently as wild-type (WT) VSV. Insertion of heterologous sequences in the juxtamembrane region completely abolished membrane fusion activity and virus infectivity, as did deletion of residues F440 to N449. The insertion mutants showed some changes in pH-dependent conformational changes and in virus binding, which could partially explain the defects in membrane fusion activity, but all the other mutants were similar to WT G with respect to conformational changes and virus binding. These data support the hypothesis that the membrane-proximal domain contributes to G-mediated membrane fusion activity, yet the conserved aromatic residues are not essential for membrane fusion or virus infectivity.

Membrane fusion activity of vesicular stomatitis virus glycoprotein G is induced by low pH but not by heat or denaturant.

The fusogenic envelope glycoprotein G of the rhabdovirus vesicular stomatitis virus (VSV) induces membrane fusion at acidic pH. At acidic pH the G protein undergoes a major structural reorganization leading to the fusogenic conformation. However, unlike other viral fusion proteins, the low-pH-induced conformational change of VSV G is completely reversible. As well, the presence of an alpha-helical coiled-coil motif required for fusion by a number of viral and cellular fusion proteins was not predicted in VSV G protein by using a number of algorithms. Results of pH dependence of the thermal stability of G protein as determined by intrinsic Trp fluorescence and circular dichroism (CD) spectroscopy show that the G protein is equally stable at neutral or acidic pH. Destabilization of G structure at neutral pH with either heat or urea did not induce membrane fusion or conformational change(s) leading to membrane fusion. Taken together, these data suggest that the mechanism of VSV G-induced fusion is distinct from the fusion mechanism of fusion proteins that involve a coiled-coil motif.

Mutations in the conserved carboxy-terminal hydrophobic region of glycoprotein gB affect infectivity of herpes simplex virus.

Glycoprotein gB is the most highly conserved glycoprotein in the herpesvirus family and plays a critical role in virus entry and fusion. Glycoprotein gB of herpes simplex virus type 1 contains a hydrophobic stretch of 69 aa near the carboxy terminus that is essential for its biological activity. To determine the role(s) of specific amino acids in the carboxy-terminal hydrophobic region, a number of amino acids were mutagenized that are highly conserved in this region within the gB homologues of the family HERPESVIRIDAE: Three conserved residues in the membrane anchor domain, namely A786, A790 and A791, as well as amino acids G743, G746, G766, G770 and P774, that are non-variant in Herpesviridae, were mutagenized. The ability of the mutant proteins to rescue the infectivity of the gB-null virus, K082, in trans was measured by a complementation assay. All of the mutant proteins formed dimers and were incorporated in virion particles produced in the complementation assay. Mutants G746N, G766N, F770S and P774L showed negligible complementation of K082, whereas mutant G743R showed a reduced activity. Virion particles containing these four mutant glycoproteins also showed a markedly reduced rate of entry compared to the wild-type. The results suggest that non-variant residues in the carboxy-terminal hydrophobic region of the gB protein may be important in virus infectivity.