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Biomedical subjects

J Morkowski

Publications and source records attributed to J Morkowski.

4 recordsLinked to original sources
PubMed1992

Rapid in vivo induction of HIV-specific CD8+ cytotoxic T lymphocytes by a 15-amino acid unmodified free peptide from the immunodominant V3-loop of GP120.

Efforts to generate a vaccine to prevent infection by human immunodeficiency virus (HIV) have focused on inducing neutralizing antibodies. However, cytotoxic T-lymphocyte (CTL) responses are a major immune defense mechanism required for recovery from many different virus infections. Since CTL epitopes can be defined by short synthetic peptides, we searched for HIV peptides that elicit a viral-specific CTL response in mice. We have developed a new method for screening CTL-inducing peptides involving a single injection into the footpad of mice to prime CTLs in the draining popliteal lymph node of mice within 10 days. Our results demonstrate that a 15-amino acid peptide (aa 315-329) derived from the V3 loop of HIV gp120 caused a rapid induction of peptide-specific and gp160-specific CD8-positive CTLs. Lysis of targets is specific since cells preincubated with unrelated peptides are resistant to lysis as are cells of a different MHC haplotype pretreated with the cognate peptide. Pretreatment of restimulated node cells with complement plus anti-CD8 but not anti-CD4 removed the lytic activity. We also successfully induced in vivo CTL activity with unmodified synthetic peptides from the influenza and Sendai virus nucleoproteins, indicating general applicability of our method for rapid screening of CTL epitopes. Because HIV replication has been reported by several labs to occur mainly in lymph nodes of infected patients, the rapid induction of HIV-specific CTLs in proximal lymph nodes by unmodified peptides emphasizes the physiological significance of our findings toward vaccine and therapeutic approaches.

Amino Acid Sequence↗
PubMed1989

Development of a monoclonal antibody specific for the gamma chain of the T-cell antigen receptor using an open reading frame expression vector.

To develop an anti-framework monoclonal antibody (mab) specific for the gamma (gamma)-chain of the T-cell antigen receptor (TCR), we expressed a part of the constant region of the gamma-chain (C gamma 2 gene segment) in E. coli using the pWR590 vector. This plasmid contains the E. coli lac promoter, operator, a truncated beta-galactosidase (beta-gal) gene (coding for the first 590 of the 1,007 amino acids of the beta-gal) and a polylinker region (at the 3' end of the beta-gal) containing nine restriction sites. These can be cleaved by any one of eight common restriction enzymes, permitting the introduction of the DNA fragment of interest. We employed the pT gamma 1 gamma-chain cDNA probe, which like the vast majority of the gamma-chain specific probes is aberrant and contains an in-frame stop codon at the junction of V and J regions. Computer analysis of the pT gamma 1 sequence revealed several MaeIII restriction sites that could result in a number of fragments. One of these fragments consisted of 245 base pairs (nucleotides 404-648) and contained most of the CI exon of the C gamma 2. Successful insertion of this fragment to the pWR590 vector was confirmed using restriction enzyme analysis. The C gamma insert was 12% of the construct. Expression of the pWR590-HpT gamma 1 recombinant plasmid in E. coli followed by SDS-PAGE analysis revealed a hybrid protein with a molecular weight of 85 kd which constituted at least 25% of the total E. coli insoluble protein. In contrast, cells transformed with the control pWR590 vector without insert expressed a 78 kd polypeptide chain. We developed several mabs against the pWR590-HpT gamma 1 hybrid protein by fusing spleen lymphocytes from BALB/c mice immunized with the pWR590-HpT gamma 1 protein, with cells of the NS1 mouse myeloma cell line. Screening of the mabs was carried out by ELISA against the pWR590-HpT gamma 1 hybrid protein and the control pWR590 beta-gal protein (beta-gal 590), derived by expressing in E. coli the pWR590 vector without gamma-chain insert. Two groups of mabs were obtained, those reacting with the pWR590-HpT gamma 1 hybrid protein only and those reacting with both the hybrid and the control beta-gal 590 proteins. The specificity of these mabs was further studied by Western blotting with similar results.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1987

C-reactive protein in management of children's burns.

Serum levels of C-reactive protein (CRP) in 48 children with scald burns were measured at 24-hour intervals. It was found that CRP levels were proportional to the area and depth of skin involved. In longitudinal studies the CRP levels were normal and admission, reached their peak at 48 hours and by 6 days had begun to fall. The response (CRP production) was increased and prolonged by infection. We conclude that sequential estimation of CRP serum levels is helpful in the management of scald burns in children.

Burns↗