Biomedical subjects

L Lin

Publications and source records attributed to L Lin.

At least 91 records · Page 5Linked to original sources

PubMed1999

Multiple mutations of the p53 gene in human mammary carcinoma.

Alteration of the p53 tumor suppressor gene is the most common genetic abnormality in human cancer. In breast cancer, depending on the stage of disease and method of detection, mutation rates of 25-60% have been observed. Multiple mutations of p53 gene in the same tumor however, are rarely reported. In this study we explored the frequency of multiple mutations of p53 gene in mammary carcinoma in a cohort of south Florida patients. Three hundred eighty-four cases of primary breast cancer diagnosed between 1984 and 1986 at the University of Miami, Jackson Medical Center were subjects of this study. Sequence analysis of exons 5 through 8 of p53 was performed on cloned PCR-amplified DNA of formalin-fixed, paraffin-embedded tumors. Two hundred thirty-four of 384 breast cancers (61%) had p53 mutation. Of those, 36 tumors showed more than one mutation; 31 tumors had two mutations, three showed three, one tumor had five mutations, and one case carried six mutations. The majority of mutations were missense (43) followed by silent (35); and most occurred within a single exon. Our study suggests that multiple mutations of p53 suppressor gene in breast cancer are more common than currently believed.

Breast Neoplasms↗

PubMed1999

The generation of nfkb2 p52: mechanism and efficiency.

nfkb2 encodes two members of the NF-kappa B/Rel family of proteins: p52 and p100. The p100 polypeptide has been proposed to serve as a precursor of p52, which corresponds to the N-terminal half of p100. While p52 functions as a Rel transcription factor, the larger p100 protein acts as a cytoplasmic inhibitor of select NF-kappa B/Rel transcription factor complexes. Because of their distinct functions, we have studied the biochemical basis for the production of these two nfkb2-derived gene products. Like the p50 product of the nfkb1 gene, p52 is principally generated in a cotranslational manner involving proteolytic processing by the proteasome. The generation of p52 is dependent on a glycine-rich region (GRR) located upstream of the p52 C-terminus, and repositioning of this GRR alters the location of proteasome processing. In most cells, small amounts of p52 are produced relative to the levels of p100, unlike the usually balanced production of nfkb1-derived p50 and p105. Using p100/p105 chimeras containing different segments of the nfkb1 and nfkb2 genes, we have found that diminished p52 processing is a property conferred by peptide sequences located downstream of the GRR, flanking the site of p52 processing.

Amino Acid Sequence↗

PubMed1999

Cognitive changes and modified processing of amyloid precursor protein in the cortical and hippocampal system after cholinergic synapse loss and muscarinic receptor activation.

A number of in vitro studies have shown that activation of muscarinic receptors by cholinergic agonists stimulates the nonamyloidogenic, alpha-secretase-processing pathway of amyloid precursor protein (APP). To determine whether increased cholinergic neurotransmission can modify the APP processing in vivo, we administered a muscarinic receptor agonist (RS86) to normal or aged rats and rats with severe basal forebrain cholinergic deficits (induced by 192 IgG-saporin). The levels of the cell-associated APP in neocortex, hippocampus, and striatum, as well as the secreted form of APP (APPs) in cerebrospinal fluid, were examined by Western blots. Additionally, we investigated the association between the altered APP levels and behavioral deficits caused by cholinergic lesions. We found that treatment with muscarinic receptor agonist resulted in decreased APP levels in neocortex and hippocampus and increased levels of APPs in cerebrospinal fluid. Regulation of APP processing by the muscarinic agonist treatment occurred not only in normal rats, but also in aged and cholinergic denervated rats that model this aspect of Alzheimer's disease. Interestingly, we found that elevation of APP in neocortex correlated with the cognitive deficits in water-maze testing of rats with cholinergic dysfunction. These data indicate that increased cholinergic neurotransmission can enhance nonamyloidogenic APP processing in intact and lesioned rats and that APP may be involved in cognitive performance.

Aging↗

PubMed1999

The sleep disorder canine narcolepsy is caused by a mutation in the hypocretin (orexin) receptor 2 gene.

Narcolepsy is a disabling sleep disorder affecting humans and animals. It is characterized by daytime sleepiness, cataplexy, and striking transitions from wakefulness into rapid eye movement (REM) sleep. In this study, we used positional cloning to identify an autosomal recessive mutation responsible for this sleep disorder in a well-established canine model. We have determined that canine narcolepsy is caused by disruption of the hypocretin (orexin) receptor 2 gene (Hcrtr2). This result identifies hypocretins as major sleep-modulating neurotransmitters and opens novel potential therapeutic approaches for narcoleptic patients.

Amino Acid Sequence↗

PubMed1999

Subcellular localization of mannose 6-phosphate glycoproteins in rat brain.

The intracellular transport of soluble lysosomal enzymes relies on the post-translational modification of N-linked oligosaccharides to generate mannose 6-phosphate (Man 6-P) residues. In most cell types the Man 6-P signal is rapidly removed after targeting of the precursor proteins from the Golgi to lysosomes via interactions with Man 6-phosphate receptors. However, in brain, the steady state proportion of lysosomal enzymes containing Man 6-P is considerably higher than in other tissues. As a first step toward understanding the mechanism and biological significance of this observation, we analyzed the subcellular localization of the rat brain Man 6-P glycoproteins by combining biochemical and morphological approaches. The brain Man 6-P glycoproteins are predominantly localized in neuronal lysosomes with no evidence for a steady state localization in nonlysosomal or prelysosomal compartments. This contrasts with the clear endosome-like localization of the low steady state proportion of mannose-6-phosphorylated lysosomal enzymes in liver. It therefore seems likely that the observed high percentage of phosphorylated species in brain is a consequence of the accumulation of lysosomal enzymes in a neuronal lysosome that does not fully dephosphorylate the Man 6-P moieties.

Animals↗

PubMed1999

Prevention of transfusion-associated graft-versus-host disease by photochemical treatment.

Photochemical treatment (PCT) with the psoralen S-59 and long wavelength ultraviolet light (UVA) inactivates high titers of contaminating viruses, bacteria, and leukocytes in human platelet concentrates. The present study evaluated the efficacy of PCT to prevent transfusion-associated graft-versus-host disease (TA-GVHD) in vivo using a well-characterized parent to F1 murine transfusion model. Recipient mice in four treatment groups were transfused with 10(8) splenic leukocytes. (1) Control group mice received syngeneic splenic leukocyte transfusions; (2) GVHD group mice received untreated allogeneic splenic leukocytes; (3) gamma radiation group mice received gamma irradiated (2,500 cGy) allogeneic splenic leukocytes; and (4) PCT group mice received allogeneic splenic leukocytes treated with 150 micromol/L S-59 and 2.1 J/cm2 UVA. Multiple biological and clinical parameters were used to monitor the development of TA-GVHD in recipient mice over a 10-week posttransfusion observation period: peripheral blood cell levels, spleen size, engraftment by donor T cells, thymic cellularity, clinical signs of TA-GVHD (weight loss, activity, posture, fur texture, skin integrity), and histologic lesions of liver, spleen, bone marrow, and skin. Mice in the control group remained healthy and free of detectable disease. Mice in the GVHD group developed clinical and histological lesions of TA-GVHD, including pancytopenia, marked splenomegaly, wasting, engraftment with donor derived T cells, and thymic hypoplasia. In contrast, mice transfused with splenic leukocytes treated with (2,500 cGy) gamma radiation or 150 micromol/L S-59 and 2.1 J/cm2 UVA remained healthy and did not develop detectable TA-GVHD. Using an in vitro T-cell proliferation assay, greater than 10(5.1) murine T cells were inactivated by PCT. Therefore, in addition to inactivating high levels of pathogenic viruses and bacteria in PC, these data indicate that PCT is an effective alternative to gamma irradiation for prevention of TA-GVHD.

Animals↗

PubMed1999

Nocturnal sleep and daytime sleepiness in normal subjects with HLA-DQB1*0602.

Narcolepsy, a neurological disorder characterized by excessive daytime sleepiness and abnormal REM sleep, is known to be tightly associated with the Human Leukocyte Antigen (HLA) DQ allele DQB1*0602. In this study, we have explored the possibility that normal subjects carrying this HLA allele (25% of the general population) could display subclinical REM sleep abnormalities and increased daytime sleepiness. Data from 525 middle-aged adults enrolled in the Wisconsin Sleep Cohort study were used for this analysis. Nocturnal polysomnography, sleep latency during the multiple sleep latency test (MSLT), and questionnaire items pertaining to excessive daytime sleepiness were compared between DQB1*0602 positive (n = 132) and negative (n = 393) participants. Results indicate shorter REM latency whether or not the latency was adjusted for wake after sleep onset (p = 0.003) and p = 0.02 respectively), increased sleep efficiency (p = 0.06) and decreased percent time spent in stage I sleep (p = 0.02) during nocturnal polysomnography in DQB1*0602 subjects. Data gathered using the Multiple Sleep Latency Test or the Epworth and Stanford sleepiness scales did not differentiate between DQB1*0602 positive and negative subjects. These results support the hypothesis that polymorphisms at the level of HLA DQ modulates sleep tendencies in humans.

Adult↗

PubMed1999

The role of bacterial vaginosis in infection after major gynecologic surgery.

PURPOSE: Previous studies have reported an association between bacterial vaginosis (BV) and postoperative fever and infection. This prospective study investigated whether the intermediate or definite stages of BV are risk factors for postoperative infection after major gynecologic surgery. METHODS: Vaginal cultures were obtained preoperatively from 175 women undergoing gynecologic surgery. The diagnostic criteria for BV were based on Nugent's standardized method of Gram stain interpretation. Postoperative fever was defined as at least one temperature equal to 101.0 degrees F or greater, or two or more temperatures more than 6 hours apart equal to 100.4 degrees F or greater. RESULTS: Thirty-six percent of the positive-BV group developed a postoperative fever, compared with 20% of the Lactobacillus-predominant group and 12% of the intermediate-BV group (P = 0.017). The differences between the positive-BV group and the Lactobacillus-predominant group, and between the positive-BV group and the intermediate-BV group, with respect to postoperative fever, were statistically significant (P = 0.045 and P = 0.007, respectively). The difference between the intermediate-BV group and the Lactobacillus-predominant group was not statistically significant (P = 0.28). CONCLUSIONS: Although the association between BV and postoperative febrile morbidity could be a spurious result of confounding with other variables, it may be prudent for the surgeon to identify patients with BV and treat them preoperatively.

Bacterial Infections↗

PubMed1999

Construction of phosphorylatable chimeric monoclonal antibody CC49 with a casein kinase I recognition site.

Phosphorylation sites for casein kinase I were introduced into chimeric monoclonal antibody CC49 (MAb-chCC49) by inserting a synthetic fragment (CK1) encoding two casein kinase I phosphorylation sites into an expression vector. The phosphorylation sites were created by incorporating the predicted consensus sequences for phosphorylation by the casein kinase I at the carboxyl terminus of the heavy-chain constant region of the MAb-chCC49. The resultant modified MAb-chCC49 (MAb-chCC49CK1) was expressed and purified. The MAb-chCC49CK1 protein can be phosphorylated by the casein kinase I with [gamma-32P]ATP to high radiospecific activity. The 32P-labeled MAb-chCC49CK1 protein binds to cells expressing TAG-72 antigens. The introduction of phosphorylation sites into MAb provides new reagents for the diagnosis and treatment of cancer. This demonstrates that, as was described for the cAMP-dependent protein kinase site, the casein kinase I recognition site can also be used to introduce phosphorylation sites into proteins.

Amino Acid Sequence↗

PubMed1999

Introduction of protein kinase recognition sites into proteins: a review of their preparation, advantages, and applications.

Labeled proteins are used in a variety of applications. This review focuses on methods that utilize genetic engineering to introduce protein kinase recognition sites into proteins. Many protein kinase recognition sites can be introduced into proteins and serve as useful tags for a variety of purposes. The introduction of protein kinase recognition sites into proteins can be achieved without modifying the essential structure or function of the proteins. Because proteins modified by these procedures retain their activity after phosphorylation, they can be used in many applications. The phosphorylatable proteins can be labeled easily to high specific activity with radioisotopes ((32)P, (33)P, or (35)S), or the nonradioactive (31)P can be used. The use of these radioisotopes provides a convenient and safe method for radiolabeling proteins. Moreover, the use of the nonradioactive (31)P with protein tyrosine kinase recognition sites permits the tagging of proteins and their detection with the many anti-phosphotyrosine antibodies available. Overall, the procedure represents a convenient, safe, and efficient method to label proteins for a variety of applications.

Amino Acid Motifs↗

PubMed1999

Dobutamine stress echocardiography in women with chest pain. Pilot phase data from the National Heart, Lung and Blood Institute Women's Ischemia Syndrome Evaluation (WISE).

OBJECTIVES: The aim of this project was to assess the utility of dobutamine stress echocardiography (DSE) for evaluation of women with suspected ischemic heart disease. BACKGROUND: Most investigations addressing efficacy of diagnosis and treatment of coronary artery disease (CAD) have been performed in predominantly male populations. As part of the Women's Ischemia Syndrome Evaluation (WISE) study, DSE was assessed in women participating at the University of Florida clinical site. METHODS: Women with chest pain or other symptoms suggestive of myocardial ischemia and clinically indicated coronary angiography were eligible for the WISE study. Enrolled subjects underwent DSE using a modified protocol. Coronary stenosis was assessed by core laboratory quantitative coronary angiography (QCA). RESULTS: The 92 women studied ranged in age from 34 to 82 years (mean 57.5). All women had > or = 1 major risk for CAD, and most (89, 97%) had > or = 2 risk factors. In 78 women (85%), left ventricular wall motion was normal at baseline and during peak infusion. The remaining 14 women had wall motion abnormalities during DSE. By QCA, 25 women (27%) had > or = 50% coronary stenosis, including 10 with single-vessel obstruction. Dobutamine stress echocardiography was abnormal in 10 of these 25 women, yielding overall sensitivity of 40%, and 60% for multivessel stenosis. Exclusion of women with inadequate heart rate response yielded overall sensitivity of 50%, and 81.8% for multivessel stenosis. Dobutamine stress echocardiography was normal in 54 of the 67 women with < 50% coronary narrowing, specificity 80.6%. CONCLUSIONS: Dobutamine stress echocardiography reliably detects multivessel stenosis in women with suspected CAD. However, DSE is usually negative in women with single-vessel stenosis, and in the larger subset without coronary stenosis. Ongoing protocols of the WISE study are expected to improve diagnostic accuracy in women with single-vessel disease, as well as provide important data in the substantial number of women with chest pain but without epicardial coronary artery stenosis.

Adult↗

PubMed1999

Monogenea of Chinese marine fishes. X. A new species and two new records of the Axinidae from fishes of the South China Sea.

This paper reports a new species and two new records of the Axinidae collected from fishes of the South China Sea. Loxuroides fungilliformis n. sp. was obtained from the gills of Hemirhamphus guoyi collected at Zhapo (21 degrees 34'N, 111 degrees 49'E). The new species is similar to L. sasikala (Unnithan, 1957) Price, 1962 in the structure of the cirrus and genital atrium, but differs from the latter in the shape and number of the spines of its genital atrium and cirrus. Both L. sasikala and Axine tripathii Price, 1962 are recorded for the first time off China.

Animals↗

PubMed1999

Characterization of three distinct cDNA clones encoding cysteine proteinases from maize (Zea mays L.) callus.

In previous work, a 33 kDa cysteine proteinase was found in callus initiated from maize (Zea mays L.) resistant to fall armyworm feeding. A callus cDNA library from the maize inbred Mp708 was screened with oligonucleotides derived from the N-terminal amino acid sequence of the 33 kDa proteinase and several cDNA clones were isolated and sequenced. A cDNA clone encoding the 33 kDa cysteine proteinase, mir1, was identified. Two additional clones, mir2 and mir3, encoding putative cysteine proteinases were also identified. mir2 and mir3 are distinct from mir1 and each other, but show a high degree of homology. All of the mir cDNA clones map to distinct sites on the maize genome. Amino acid sequences encoded by the mir clones are similar to other known cysteine proteinases and are most closely related to the oryzain-alpha and -beta precursors. The ERFNIN motif and a 12 amino acid conserved sequence are present in the propeptide region of the putative proteinases encoded by mir clones. mir2 and mir3 appear to have C-terminal extensions. The phylogenetic tree of nucleotide sequences of mir1, mir2, mir3 and other representative cysteine proteinases from protozoa, plants and animals was constructed.

Amino Acid Sequence↗

PubMed1999

Localization of the N-methyl-D-aspartate R1 receptor in the pituitary gland of immature rainbow trout (Oncorhynchus mykiss).

A study was conducted on the selective immunostaining of pituitary cells and pars nervosa of immature rainbow trout by antibody raised against the R1 subunit of the rat ionotropic N-methyl-D-aspartate (NMDA) receptor. In the pars distalis, the mammotrophs of the rostral pars distalis exhibited the most marked imunoreactivity, with the somatotrophs of the proximal pars distalis showing a consistently lower degree of immunoreactivity. No imunoreactivity was associated with the corticotrophs, thyrotrophs or gonadotrophs. In the pars intermedia, the melanotrophs showed no evidence of immunoreactivity, whereas the putative somatomammotrophs exhibited a wide inter-animal range of immunoreactivity. Some imunostaining was also evident in the anterior and posterior part of the pars nervosa, at the interface of the pars distalis or pars intermedia with the pars nervosa.

Animals↗

PubMed1999

Compound heterozygosity for novel splice site mutations in the BPAG2/COL17A1 gene underlies generalized atrophic benign epidermolysis bullosa.

Generalized atrophic benign epidermolysis bullosa, GABEB (OMIM# 226650), is a nonlethal variant of epidermolysis bullosa with autosomal recessive inheritance pattern. The pathogenesis of this disorder can be caused by mutations affecting two different gene/protein systems. Most of the mutations have been identified in the BPAG2/COL17A1 gene encoding a hemidesmosomal transmembrane protein, the 180 kDa bullous pemphigoid antigen (BP180), also known as type XVII collagen. The minority of the mutations are localized in the LAMB3 gene encoding the beta3 polypeptide of laminin 5. In In this study we describe a GABEB patient who showed absent expression of BP180 in the cultured keratinocytes as well as in the skin. The patient was a compound heterozygote for two different splice site mutations, 3053-1G-->C and 3871+1G-->C, affecting the extra-cellular domain of the protein. These mutations resulted in multiple aberrant splice variants, three of them causing premature termination codons for translation. This case, dealing with out-of-frame splice site mutations in BPAG2/COL17A1, attests to the molecular heterogeneity of GABEB.

Adult↗

PubMed1999

Up-regulation of hepatocyte growth factor receptor: an amplification and targeting mechanism for hepatocyte growth factor action in acute renal failure.

BACKGROUND: Hepatocyte growth factor (HGF) and its c-met receptor comprise a signaling system that has been implicated in tissue repair and regeneration. HGF action is specifically targeted to the damaged organ following injury; however, the mechanism underlying this important targeting process remains to be elucidated. We reasoned that induction of c-met expression might be a critical factor in determining the site specificity of this receptor-ligand system. To test this hypothesis, we examined changes in activity of the HGF/c-met system in the folic acid model of acute tubular injury and repair. METHODS: Tissue HGF and c-met mRNA levels were detected by RNase protection assay and Northern blot analysis following acute renal injury induced by a single injection of folic acid. HGF and c-met proteins were examined by a specific enzyme immunoassay and Western blotting, respectively. C-met expression and trans-activation were investigated by exposing renal epithelial mIMCD-3 cells to various cytokines in vitro. RESULTS: Extremely rapid induction of renal HGF and c-met mRNA was observed beginning one hour following injection of folic acid. Circulating plasma HGF protein level rose dramatically (approximately 16-fold), peaking first at two hours and again at 24 hours following injection. Despite elevated HGF mRNA in the kidney, total kidney HGF protein actually decreased significantly at 24 hours following injury. On the other hand, both c-met mRNA and c-met protein were markedly increased in the kidney, where active renal tubule repair and regeneration take place. In vitro studies suggested that increased levels of HGF, as well as other cytokines, might account for enhanced c-met expression in renal tubular epithelial cells. Pretreatment of the cells with actinomycin D totally blocked c-met induction, suggesting that induced c-met expression occurs primarily at the transcriptional level. Using a cloned region of the c-met promoter coupled to a reporter gene, we demonstrated that HGF directly stimulated c-met promoter transactivation in renal epithelial cells. CONCLUSION: These results suggest that local up-regulation of c-met transcription in the kidney is crucial to renal tubule repair and regeneration, not only because it increases overall activity of this receptor-ligand system, but also as a mechanism targeting HGF action specifically to renal epithelia.

Acute Kidney Injury↗

PubMed1999

Elimination of cytokine production in stored platelet concentrate aliquots by photochemical treatment with psoralen plus ultraviolet A light.

BACKGROUND: Cytokines generated in platelet concentrates (PCs) during storage have been implicated as possible mediators of febrile nonhemolytic transfusion reactions. Two potential methods of white cell inactivation were compared for their ability to reduce cytokine synthesis in pooled random-donor PC aliquots: treatment with gamma-radiation and photochemical treatment (PCT) using psoralens and ultraviolet A light. STUDY DESIGN AND METHODS: ABO-matched PC aliquots were pooled and divided into separate aliquots. Aliquots (20 mL) were taken from each pool to serve as an untreated control and to undergo gamma-radiation. Aliquots were treated by using either gamma-radiation (2500 or 5000 cGy) or virucidal PCT. PCT with the psoralens 8-methoxypsoralen (8-MOP), aminomethyltrimethyl psoralen (AMT), and S-59 was investigated. PC aliquots were stored for 7 days and analyzed for levels of interleukin 8 by use of an enzyme-linked immunosorbent assay. Levels of DNA adduct formation were determined by using 3H-labeled psoralens. RESULTS: Levels of interleukin 8 in the untreated random-donor PC aliquots increased with increasing white cell counts, but they were not affected by pooling. The untreated control aliquots and the aliquots treated with gamma-radiation had significant increases in levels of interleukin 8 after 5 to 7 days of storage (p<0.05). PCT with S-59 resulted in a significant reduction in cytokine synthesis (p<0.05). Day 5 to 7 levels of interleukin 8 did not differ significantly from Day 0 levels. Inhibition of interleukin 8 production by PCT increased with increasing levels of DNA modification (S-59 > AMT > 8-MOP). CONCLUSION: PCT that utilizes S-59 has been developed to inactivate potential viral and bacterial pathogens in PC aliquots while maintaining in vitro platelet function. These data demonstrate that PCT of aliquots of pooled PC aliquots before storage also prevents white cell cytokine synthesis during storage. PCT may therefore offer the potential for reducing cytokine-associated febrile nonhemolytic transfusion reactions.

Blood Preservation↗

PubMed1999

Expression of endogenous retrovirus ERV-3 induces differentiation in BeWo, a choriocarcinoma model of human placental trophoblast.

Differentiation-related expression of endogenous retrovirus ERV-3 env in the normal human placental syncytiotrophoblast suggests a role in placental development. The choriocarcinoma cell line BeWo, a model of trophoblast differentiation, is maintained in an undifferentiated state and undergoes differentiation upon the addition of forskolin. The expression of ERV-3 env mRNA increased after 48 h forskolin treatment, concurrently with increased intercellular fusion and production of human chorionic gonadotropin (beta-hCG) mRNA, a hormonal differentiation marker for trophoblast. Over expression of ERV-3 env induced differentiation of BeWo characterized by decreased cell growth, differentiation-related morphologic changes, and induction of beta-hCG mRNA. These results support the first known role for the expression of an endogenous retrovirus in trophoblast differentiation.

Cell Differentiation↗