Immunological and molecular analysis of the sentinel lymph node: a potential approach to predict outcome, tailor therapy, and optimize parameters for tumor vaccine development.
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Animals↗
Biomedical subjects
W J Wood
Publications and source records attributed to W J Wood.
Examining the immune response in sentinel lymph nodes of mice and men.
Recently, it was recognized that an immune response develops along one of two major pathways. One leads to a destructive immune response (type 1), while the alternative leads to a nondestructive immune response (type 2). Our studies in animal models suggest that therapeutic vaccines induce a tumor-specific type 1 immune response while ineffective vaccines induce a type 2 response. These results have led us to examine the immune response in sentinel lymph nodes draining tumor vaccines of patients entered onto clinical trials for melanoma, breast and renal cell cancer.
Animals↗
Alternatively spliced exons encode the tissue-specific 5' termini of leukocyte pp52 and stromal cell S37 mRNA isoforms.
The pp52 gene encodes an intracellular, F-actin-binding phosphoprotein (also designated LSP1 and WP34) postulated to function in cytoskeleton dynamics and cell motility. We previously reported that different mRNA isoforms are expressed from this gene in cells of the leukocyte lineage versus mesodermally derived cells. These tissue-specific mRNA isoforms are identical except for 5'-untranslated regions and sequences coding for unique N-termini of 23 and 21 amino acids, respectively. As this is a single-copy gene, we predicted that these tissue-specific mRNA isoforms would be generated by alternative RNA splicing. We report that the unique 5' sequences in these mRNA isoforms are encoded in two separate exons containing ATG initiation codons. These features confirm that the pp52 and S37 mRNA isoforms are generated by alternative RNA splicing and establish that they are independently translated. Other results presented here indicate that the differential expression of these exons in leukocytes versus mesodermally derived cells is regulated at the level of transcription by tissue-specific promoters.
Alternative Splicing↗
The promoter and 5' flanking sequences controlling human B29 gene expression.
The product of the B-cell-specific B29 gene (B29, Ig beta, CD79b) is essential for Ig-mediated B-cell activation via the B-cell antigen receptor complex (BCR) on human and murine B lymphocytes. To better understand the regulation of this pivotal gene, we have analyzed the human genomic DNA sequence upstream of the B29 ATG start codon for transcriptional control activity. The human B29 gene lacks either a TATA or a CAAT box and transcription is initiated at multiple sites. The minimal promoter of the human B29 gene is contained within a 193-bp region 5' of these multiple start sites. This minimal promoter exhibits B-cell-specific activity and contains SP1, ETS, OCT, and IKAROS/LYF-1 transcription factor motifs. All these motifs are strikingly conserved in sequence and placement relative to the previously characterized murine B29 promoter. Additional upstream gene segments dramatically affected B29 minimal promoter activity. A newly identified motif called the B29 conserved sequence (BCS), found upstream of both human and murine B29 promoters, appears to stimulate B29 transcription through a novel mechanism. A single BCS had little effect either on the minimal B29 promoter or on a heterologous promoter. Instead, the BCS stimulated transcription by counteracting 5' negative regulatory DNA sequences that block the activity of the B29 minimal promoter in its absence. These findings indicate that B29 gene expression is controlled by the complex interplay of positive and negative regulatory elements.
Amino Acid Sequence↗
Long-term results of laser treatment in the ocular histoplasmosis syndrome.
OBJECTIVE: To determine the long-term visual outcome, rate of persistent choroidal neovascularization, and rate of recurrent choroidal neovascularization in eyes undergoing laser photocoagulation for choroidal neovascularization secondary to ocular histoplasmosis syndrome. DESIGN AND PATIENTS: One hundred one eyes with 5 to 16 years of follow-up that presented with choroidal neovascularization secondary to ocular histoplasmosis were retrospectively evaluated. Patients were grouped according to location of choroidal neovascularization and assignment to observation or laser photocoagulation. MAIN OUTCOME MEASURES: Visual acuity outcome and loss for all groups were compared. The rates of persistent and recurrent choroidal neovascularization for the treated eyes were also evaluated. RESULTS: Visual acuity of 20/40 or better was obtained in 71% of eyes with treated extrafoveal choroidal neovascularization and 68% with treated juxtafoveal choroidal neovascularization. Recurrent choroidal neovascularization was observed in 23% of treated eyes during a mean follow-up of 9.6 years. CONCLUSION: Results support the long-term benefit of photocoagulation and need for careful follow-up.
Adolescent↗
Short-term night blindness associated with colon resection and hypovitaminosis A.
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Colectomy↗
Multiple B29 containing complexes on murine B lymphocytes. Common and stage-restricted Ig-associated polypeptide chains.
The B29 gene is specifically expressed in all cells of the B lymphocyte lineage, and the B29 protein is disulfide-linked to the protein product of at least one other gene, known as mb-1. The noncovalent association of these heterodimers with Ig H chains is thought to be required for surface expression and signal transmission by B cell Ag receptors. We now demonstrate by two-color immunofluorescence a direct correlation between B29 density and surface Ig expression on normal spleen and bone marrow cells. The proportion of B29 in Ag receptor complexes appears to be relatively constant across major B lymphocyte subpopulations. Multiple B29-containing heterodimers were resolved on normal spleen cells by surface labeling, immunoprecipitation, two-dimensional gel analysis, and immunoblotting. As with lymphoma cells in our earlier study, the conditions of detergent extraction were critical to detection of certain species. Many laboratories have observed a family of 69- to 85-kDa heterodimers that are extracted with digitonin. These species are clearly Ig-associated, and are coprecipitated with anti-Ig antibodies. We found that extraction with Triton X-100 revealed an additional pair of 52- to 58-kDa heterodimers, where B29 was disulfide-bonded to a protein of approximately 23 kDa. The latter was detectable by immunoblotting with antibodies to extracellular, but not cytoplasmic, portions of mb-1. We found that, with mature cells, both conventional and low molecular mass heterodimers were solubilized with digitonin, but only detectable if Triton was present during immunoprecipitation. Thus, a protein having partial serologic identity with mb-1 forms heterodimers that are cryptic on splenic B cells, and possibly not directly associated with surface Ig molecules. In contrast, both types of heterodimers were readily detectable on late stage pre-B cells, regardless of detergent used for extraction or antibody used for immunoprecipitation. In that situation, both low- and high molecular mass heterodimers were associated with surface Ig. These findings increase our understanding of the B lymphocyte Ag receptor complex and indicate that its components may change as a function of differentiation.
Animals↗
Alternatively spliced pp52 mRNA in nonlymphoid stromal cells.
The 52-kDa phosphoprotein, also reported as lymphocyte-specific gene 1 and WP34, is transcribed as a 1.6-kb mRNA in B lymphocytes, B cell lines, and untransformed T cells. This gene encodes a cytoplasmic and plasma membrane-associated protein that is phosphorylated at a casein kinase II site and reportedly binds calcium. Based on these properties, it has been hypothesized that lymphoid form of the 52-kDa phosphoprotein protein may play a role in lymphocyte signal transduction. We show that alternatively spliced mRNA are expressed from this gene in nonlymphoid cell lines (myocytes, stromal cells, fibroblasts). These cell lines do not express the 1.6-kb lymphoid cell-specific transcript. Instead, mRNA of 2.0 and 2.8 kb are detected in varying abundance. A full-length 2.0-kb cDNA has been cloned and sequenced from the BMS2 stromal cell line by conventional screening and polymerase chain reaction-based methods. This cDNA clone, designated S37, has a single open reading frame encoding a 328 amino acid peptide. The nucleotide sequence of the S37 stromal cell cDNA is identical to that of the lymphocyte derived pp52 cDNA from the 3' poly(A) tail to the codon encoding the amino acid at residue 24. This region of the S37 cDNA clone encodes a protein that is identical to that encoded by the lymphoid pp52 cDNA and includes a casein kinase II phosphorylation site. However, the two clones differ in their 5' nucleotide sequence and their NH3 terminal amino acid sequence. This organization is consistent with alternative exon utilization. These results suggest that tissue-specific control mechanisms are used to generate different forms of lymphoid form of the 52-kDa phosphoprotein mRNA in lymphoid cells versus mesoderm-derived, nonlymphoid cell lineages.
Alternative Splicing↗
Laser treatment for subfoveal neovascular membranes in ocular histoplasmosis syndrome: results of a pilot randomized clinical trial.
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Choroid↗
Human lymphocyte-specific pp52 gene is a member of a highly conserved dispersed family.
For a better understanding of genes that potentially function in B lymphocyte cell signaling, we isolated the human genomic counterpart of the murine pp52 or LSP1 gene. We unexpectedly found that the human pp52 gene is one of four closely related loci. Representative cosmids from each of the four family members were isolated and chromosomally localized by fluorescence in situ hybridization. Nucleotide sequence was obtained from an exon common to each locus and demonstrated very close similarity among all four loci. Two of the four loci harbored dysfunctional frameshift mutations or premature translation stop sites. The exon of one locus was flanked by an 80-bp perfect inverted repeat, suggesting that it may have originated through a looped intermediate DNA structure. Through a series of cDNA hybridization studies and nucleotide sequence analyses we were able to unambiguously link the lymphocyte-expressed gene to the locus mapped to chromosome 11p15.5. This same chromosomal band has been involved in tumor-related chromosomal translocations found in chronic lymphocytic leukemia.
Animals↗
Isolation and chromosomal mapping of the human immunoglobulin-associated B29 gene (IGB).
The B29 gene encodes a B-cell-specific membrane protein in the immunoglobulin antigen receptor complex. B29 is a crucial member of this receptor complex and is believed to function as an effector of signal transduction in a manner analogous to that of the CD3 components of the T cell antigen receptor. We have isolated a full-length human B29 cDNA clone by using a murine B29 cDNA probe. We show that there is an extremely high degree of evolutionary conservation between the human and mouse proteins, particularly in the transmembrane and intracytoplasmic regions, where the identity is 96%. In addition, the intracytoplasmic region in both proteins contains an identical peptide motif that is present in a number of molecules involved in lymphocyte activation. Genomic Southern blot analysis of human cell lines hybridized with both murine and human B29 cDNAs gives patterns consistent with a single-copy gene occupying a small region of the genomic sequence. Using human B29 cosmid DNA, we have localized the B29 gene to human chromosome 17q23 via fluorescence in situ hybridization. B29 is the first gene localized to this area of the genome. Interestingly, a subset of human B cell chronic lymphocytic leukemias (CLL) has translocations in this locus on chromosome 17.
Amino Acid Sequence↗
B29 gene products complex with immunoglobulins on B lymphocytes.
B29 is a B-lineage-specific gene predicted from sequence information to be a transmembrane member of the immunoglobulin (Ig) superfamily, with a single extracellular Ig-like domain. Its presumptive cytoplasmic region contains a peptide motif present in CD3 and other molecules involved in lymphocyte activation. Affinity-purified goat antibodies were prepared to a TrpE fusion protein of B29 and used to study B29 expression on lymphoid cells. The antiserum precipitated surface-labeled heterodimers from B lymphoma cells. One was 65-88 kDa (unreduced) or 36-47 plus 32-34 kDa (reduced) by SDS/PAGE analysis, regardless of detergent. A smaller heterodimer was detected only with Triton detergent extraction. IgM molecules were coprecipitated by the B29 antiserum when the weak detergent digitonin was used. In addition, cocapping experiments revealed that most B29 molecules codistribute with Ig on the cell surface. Although early B-lineage cells and plasma cells contain B29 mRNA, surface expression was detectable only on B cells that had significant amounts of surface Ig. The surface expression was B-lineage-specific and included cells from mutant xid mice and B-cell lines representing mu, delta, gamma, and alpha heavy-chain isotypes and both kappa and lambda light-chain types. The density of surface B29 protein correlated directly with surface mu heavy-chain density on subclones of a B-cell lymphoma and lipopolysaccharide-stimulated pre-B cells. These findings show that B29 is covalently linked in a heterodimer and are consistent with a recently proposed model of surface Ig complexes.
Animals↗
The stairway to recovery. An emerging worldview uncovers God's wisdom in nature.
What is lacking for the renewal of the earth is not so much ethical consensus as political will. The myth of "man" as the self-sufficient and autonomous master of "inanimate" nature has been ingrained for centuries, and it will take a powerful force to dislodge it. However, we are in a unique period of convergence--a time when we might be transformed by the truth if we only seek it. This new spiritual awakening represents a conscious commitment to a common cause that lies beyond individual self-interest. The commitment has been kindled by a perception that we are creatures intimately connected with the rest of creation and to the power that has called us into being. And it is a commitment carried out in action through a disciplined way of life. If we want to heal the earth, we have to get better ourselves, because it is the way we think and live and act that is harming the earth. We have much to learn from participants in Alcoholics Anonymous, who have learned the secret of proclaiming the good news by doing it, one step at a time, by living it, day by day. The first step is acknowledgment that, as a society--indeed, as the human race-we are at the bottom and cannot get back up. Once we admit we have hit the bottom, there is hope for the environment because we have taken the first step to recovery.
Alcoholics Anonymous↗
Diagnosis of bacterial vaginosis in a routine diagnostic laboratory.
Vaginal swabs from 299 hospital and general practitioners' patients were examined for Gardnerella vaginalis by Gram film and by culture, G. vaginalis was isolated in 12% of cultures. Comparison between the 'clue' cell and culture methods suggested that the former is a rapid, acceptable routine screening method for the detection of G. vaginalis. The value of the traditional method of identifying G. vaginalis by sensitivity testing is questioned. All specimens were also examined by Gram film and culture, for the presence of Mobiluncus spp, which was detected in 8.4% of specimens by Gram film but only 0.7% by culture. From a questionnaire returned by 84% of clinicians, metronidazole was found to be the most commonly used antimicrobial agent for the treatment of G. vaginalis, and in all but one case appeared to be clinically effective.
Bacteria, Anaerobic↗
Idiopathic sclerochoroidal calcification.
Idiopathic sclerochoroidal calcification is a disorder that is often overlooked, occurring at the level of the posterior sclera and choroid in older patients. Clinically, it may be confused with choroidal osteoma, choroidal metastasis, amelanotic choroidal nevus, or choroiditis. The authors present seven cases of this unusual entity. The patients ranged in age from 58 to 80 years. In all cases, the yellow lesions were clinically estimated to be at the level of the choroid and sclera, and they were usually located superior to the superotemporal retinal vascular arcade. Systemic investigation of calcium and phosphate metabolism was normal in all patients. The classic clinical manifestations and differentiation of this unusual entity from other similar fundus lesions is discussed.
Aged↗
Adriamycin-induced resistance to natural killer (NK)-mediated cytotoxicity.
Studies were done to determine the susceptibility of a colon carcinoma cell line, LoVo, to natural killer (NK) mediated lysis after exposure to the tumor cells to Adriamycin (ADR) (doxorubicin; Adria Laboratories, Columbus, OH). LoVo cells were exposed to ADR (0.4 micrograms/ml) for various time intervals and then tested for sensitivity to lysis by NK cells from the peripheral blood of normal donors in a sodium chromate (Cr51) release cytotoxicity assay. Exposure of tumor targets to ADR induced a resistance to NK-mediated lysis. Susceptibility to lysis decreased progressively to approximately 60% of control levels after 72 hours of ADR exposure. The induction of resistance was dependent on ADR dose, but did not magnify with doses greater than 0.4 micrograms/ml. When target cells were allowed to recover from ADR in fresh medium for 48 hours, complete reversal of the ADR effect was seen. The effect of interleukin-2 (IL-2) stimulation on the NK lysis of LoVo also was tested. IL-2-stimulated effector cells demonstrated enhanced cytotoxicity to LoVo targets and were able to overcome the ADR-induced resistance to lysis. The mechanism of resistance does not appear to be related to the altered binding of effectors to chemotherapy-treated targets, as suggested by single cell assay results.
Cell Differentiation↗
Current results of treatment for distant metastatic well-differentiated thyroid carcinoma.
Between 1960 and 1986, 37 patients were treated at M. D. Anderson Hospital and Tumor Institute, Houston, Tex, for metastases beyond the thyrocervical region arising from well-differentiated thyroid carcinomas. Patient, tumor, and treatment factors were analyzed in this group for their influence on survival. No specific factors achieved statistical significance; however, patients having more than one site of metastasis and patients whose metastases arose less than 5 years after treatment of the primary tumor tended to have shorter overall survival times. More than one modality of treatment was used in the majority of patients. Complete surgical resection of metastases offered the best chance for prolonged survival and palliation. With long survival times being the rule, several treatment modalities are often required to provide continuous palliation.
Adenocarcinoma↗
Hepatocellular carcinoma: importance of histologic classification as a prognostic factor.
The characteristics and clinical courses of 15 patients with fibrolamellar carcinoma have been compared to 62 patients with hepatocellular carcinoma treated over the same time period. Marked differences were found in patient, tumor, and treatment characteristics. Survival rates in these two groups of patients have been compared by univariate analysis of the subsets of those patient, tumor, and treatment characteristics. The longer survival of the group of patients with fibrolamellar carcinoma could not be accounted for by the prevalence or absence of any of these characteristics. Comparison of subsets of patients with documented noncirrhotic livers or those having complete resection of their tumors revealed a significantly prolonged survival in patients with the fibrolamellar variant. The 5 year survival rate of the patients with fibrolamellar carcinoma who had complete tumor resection was 45 percent, with a median survival time of 50 months. The 5 year survival of patients with hepatocellular carcinoma who had complete tumor resection was 0 percent, with a median survival time of 22 months.
Adolescent↗