Iranian Biomedical Journal
Volume:1 Issue: 1, Jan 1997

Human immunodeficiency virus (HIV) transactivator Tat is a potent activator of both viral and cellu‌lar genes. Tat has also been implicated in the development of AIDS-related malignancy. Here, we show that Tat physically and functionally is able to sequester the cell cycle check point protein p53. This sequestration results in non-functional promoter activity of cyclin-dependent kinase/cyclin in‌hibitor, namely p21 (Waf1). Therefore, it is proposed that a Tat/p53 complex in vivo may deregulate p53 responsive genes, hence allowing for deregulation of check point, which may lead to development of malignancies.

A new strategy for construction of synthetic gene encoding human basic fibroblast growth factor comprising DNA annealing-ligation and augmentation by polymerase chain reaction was introduced. The sequence of the gene and corresponding amino acid chain were modified in order to increase stability of the protein. First, 300 bp and 160 bp fragments of the gene were assembled from 18 oligonucleotides and ligated separately. Then, the shorter fragment was completed by using PCR and combined with the longer one in a proper orientation in pUC 19. One extra nucleotide that had been found in the gene after DNA sequencing and resulted in frame shift, was rectified through the use of PCR directed mutagenesis. Finally, 5'-terminal region of the gene was augmented by means of PCR in order to restore the N-terminal part of the protein and to introduce the NdeI recognition site. The gene was subcloned into the inducible pET-3a expression vector under control of T7 promoter and expressed in Escherichia coli. The identity of the recombinant protein and level of expression were detected by using Western blot analysis and immunoassay. The proposed method has provided a useful strategy for synthesizing modified proteins that might be applied for protein engineering.

The Hepatitis B Surface antigen (HBsAg) gene was isolated from an Iranian HBeAg positive carrier by PCR. The gene was cloned in pUC19 for sequencing and pYES2 for expression in Saccharomyces cerevisiae, which pNF1 and pDF3 constructs were made respectively. The sequencing data showed that the isolated HBsAg gene shared more than 90% homology with the ayw subtype. The pDF3 was transferred into the yeast strain and expression of the HBsAg was induced. Yeast-derived HBsAg was purified by immunoaffinity chromatography and checked by ELISA, Western blot analysis and con‌firmatory ELISA. The immunogenicity test in mice showed that injection of the yeast-derived re‌combinant HBsAg could induce the immunologic response against the antigen. 320 fold dilution of the mouse serum was positive for anti-HBsAg. The amount of HBsAg expressed in yeast constitute 1% of the total cell soluble protein.

The relationship between the immunoglobulin (Ig) nucleotide sequence and the ability of a B cell to develop into a malignant cell was studied in a subset of B cells, B-1 cells. B-1 cells become malignant in chronic lymphocytic leukemia (CLL) and are responsible for the production of "natural autoanti‌bodies". The autoimmune NZB mouse has been known as a human malignancy and CLL model, be‌cause of the age-dependent onset of clonally expanded hyperdiploid B-1 cells in these mice. The Ig heavy chain variable region in hyperdiploid B-1 clones from several NZB mice showed common char‌acteristics in the CDR3 shared with fetal B cells: lack of N base insertions and presence of homology sequences at the VH-D-JH junctions that can be encoded by either of the two joined gene segments. Using a degenerative oligoprimer was shown no significant differences in expression of the restricted CDR3/DFL16 region in newborns or in the liver of either strain of mice as young adults. However, the expression of the restricted CDR3/DFL16 in the spleens of young adult NZB was remarkably ele‌vated and showed significant differences from the expression in newborn NZB as well as from young adult and newborn BALB/c mice. It appears that malignant hyperdiploid B-1 cells are derived from fetal B cells. This technique can be used as a molecular marker to demonstrate a relative increase in the expression of this CDR3 in animals pre-destined to develop B-malignancies.

Compared to other methods of vaccine production, (e.g. culture flask and bioreactor system), the roller culture method for producing rabies vaccine is simpler, and the equipment easier to install and maintain, and also in the event of contamination of one roller, the entire batch need not be discarded. This system provides a much larger surface area than the culture flask, and also ensures a much higher surface area to volume ratio. Gas exchange is also made easier because the cells are alternately aerated and fed with medium every few minutes. Production of rabies vaccine using BHK-21 culti‌vated by roller bottle cell culture technique can be suitable for countries where rabies is an important health problem. For this project, the PV-PARIS/BITK-21 rabies virus strain was used. The vaccine thus obtained was tested on mice and a satisfactory immune response was observed, and also the po‌tency of vaccine met the requirements of the World Health Organization (WHO).

Delayed-type hypersensitivity reaction measured by skin testing, is an important tool for evaluation of cellular immune response in leishmaniasis and has been used recently as an indication of exposure to infection. Skin testing in leishmania infection needs a standard reagent with high specificity, sen sitivity, and potency. In the present work, a soluble leishmanin, and three whole parasite prepara tions from Leishmania major, i.e. thimerosal, phenol, and autoclaved leishmanins, were prepared un der standard conditions. These antigens were evaluated and compared in different foci of leishmani asis. The sensitivity and potency of reagents were tested in both recovered cases and patients with ac tive ulcer in two areas endemic to urban and rural leishmaniasis. Data obtained showed that, the sensitivity of thimerosal, phenol, and soluble leishmanins are almost equal, but are higher than auto claved one. Moreover, the potency of soluble leishmanin was also proved to be higher than other reagents. The purity of soluble leishmanin is higher than other three particulate preparations, and it lacks the majority of membrane antigens. As a result this reagent can be used as a standard and ideal antigen for skin testing in human leishmaniasis.

Enterobius vermicularis is a nematod of large intestine in man. Extra intestinal localization of this parasite is rare. A 60 year old man was admitted to the hospital with the clinical manifestation of meningitis. In laboratory examination of cerebrospinal fluid (C.S.F), the appearance was red, Albu-min level was high and the glucose showed normal value. Haematoxylin and Eosin stained slides pre‌pared from the precipitation revealed adults, larvae and ova of Enterobius vermicularis. This is the first report of Enterobius vermicularis in cerebrospinal fluid.

Among the female sexual hormones, the estrogen and progesterone are of significant importance. These hormones are prescribed for the treatment of certain female genital system disorders as well as used in the synthesis of contraceptive drugs. It has been shown that administration of the above hor‌mones during early pregnancy may produce abortion in early gestational days. In the present investi‌gation, we have studied the possible abortional and teratogenical effects of a single intramuscular injection of 0.15 mg/kg esteradiol valerate during the first or second half of pregnancy, in rats. The data show that (1) The abortional effects of administration of esteradiol during the first half of pregnancy, in comparison with the control group, is significant. Although the injection of this hor‌mone during the second half of pregnancy does not produce total abortion but there is a reduction in the number of fetuses. Therefore, it may be said that with injection of a single dose of hormone dur‌ing the first half of pregnancy (from day zero to day 10) the chance of abortion is high (in human embryos, this period is about the first four weeks of development). (2) Administration of esteradiol may also produce severe fetal abnormalities such as anencephalia, exencephalia, microphthalmia. In addition to the above abnormalities, in the male fetuses, there is also a remarkable defect in the uro‌genital system.

The gene encoding alpha fetoprotein (locus symbol Afp) was assigned to rat chromosome 14 at band p21-p22 using fluorescence in situ hybridization method. The present result suggests that there is a conserved syntenic group between human 4q11-q13, mouse 5F-G, and rat 14p21-p22.

The main aim of this study was to find a suitable rabies vaccination protocol which would allow usage of lower volumes of vaccine, fewer clinical visits and above all, enough serological effectiveness for high-risk individuals requiring pre-exposure immunization. Human diploid cell rabies vaccine was administered intradermally and intramuscularly in 60 previously unvaccinated volunteers, aged 19 to 21, following different vaccination protocols. The participants were divided into six groups, and ra‌bies antibody titration was accomplished by ELISA. The comparison of the mean titers 30 days after termination of vaccination protocols shows that a protocol requiring only two clinical visits and ID injection of a total volume of only 0.3 ml of human diploid cell rabies vaccine (0.1 + 0.1 ml on day 0, one in each arm, and 0.1 ml on day 28) provides good serological response (6.92 IU/m1).

Over a Period of three months (summer), 108 samples of water collected from four hospitals in Te-hran. They were taken from numerous potable and non-potable water, including main supply cis-terns, calorifiers, hot and cold tap water, shower, cooling tower and drain of air condition system. Culturing of concentrated and non-concentrated water samples on Buffered Charcoal Yeast Extract (BCYE) Agar and Selective BCYE agar yielded one strain of Legionella pneumophila. It was found in cooling tower drain of one hospitals. The strain isolated belonged to sero-group 4 (Portland-1 Strain). It was also atypical being hippurate negative.