Biomedical subjects
W Liu
Publications and source records attributed to W Liu.
Alanine scan of endothelin: importance of aromatic residues.
A systematic approach to map the functional important determinants of endothelin-1 (ET) by an alanine scan is described. Studies on the in vitro receptor binding affinity and on the agonist contracting activity defined that residues Asp8, Tyr13, Phe14, Leu17, and Trp21 were of major biological significance. A striking observation was that four out of these five sites were hydrophobic amino acids. Ala analogues of the aromatic residues at position 13, 14, and 21 displayed sharply reduced receptor binding affinity (< 2% of ET) and can be considered important for receptor contact. Ala analogues of Asp8 and Leu17 lost most (> 90%) of the agonist activity but retained a receptor affinity nearly equivalent to ET and can be considered to be important for signal transduction. Three other positions, Val12, Asp18, and Ile20 (which are adjacent to the biologically important sites of Tyr13, Leu17, and Trp21), resulted as partially tolerant to Ala substitution, retaining 14-50% of the potency of ET. Ala analogues of the Et isomeric disulfide arrangement (Cys1,11 and Cys3,15) were always less active than the corresponding analogues with the native disulfide pairings (Cys1,15 and Cys3,11).
Enhanced stability and potency of novel growth hormone-releasing factor (GRF) analogues derived from rodent and human GRF sequences.
Native human GRF(1-44)-NH2(hGRF44) is subject to biological inactivation by both enzymatic and chemical routes. In plasma, hGRF44 is rapidly degraded via dipeptidylpeptidase IV (DPP-IV) cleavage between residues Ala2 and Asp3. The hGRF44 is also subject to chemical rearrangement (Asn8-->Asp8, beta-Asp8 via aminosuccinimide formation) and oxidation [Met27-->Met(O)27] in aqueous environments, greatly reducing its bioactivity. It is therefore advantageous to develop long-acting GRF analogues using specific amino acid replacements at the amino-terminus (to prevent enzymatic degradation): residue 8 (to reduce isomerization) and residue 27 (to prevent oxidation). Inclusion of Ala15 substitution (for Gly15), previously demonstrated to enhance receptor binding affinity, would be predicted to improve GRF analogue potency. Substitution of [His1,Val2]-(from the mouse GRF sequence) for [Tyr1,Ala2]-(human sequence) in [Ala15,Leu27]hGRF(1-32)-OH analogues completely inhibited (24-h incubation) DPP-IV cleavage and greatly increased plasma stability in vitro. Additional substitution of Thr8 (mouse GRF sequence), Ser8 (rat GRF sequence), or Gln8 (not naturally occurring) for Asn8 (human GRF sequence) resulted in analogues with enhanced aqueous stability in vitro (i.e., decreased rate of isomerization). These three highly stable and enzymatically resistant hGRF(1-32)-OH analogues, containing His1, Val2, Thr/Gln8, Ala15, and Leu27 replacements, were then bioassayed for growth hormone (GH)-releasing activity in vitro (rat pituitary cell culture) and in vivo (SC injection into pigs). Enhanced bioactivity was observed with all three hGRF(1-32)-OH analogues. In vitro, these analogues were approximately threefold more potent than hGRF44, whereas in vivo they were eleven- to thirteenfold more potent.(ABSTRACT TRUNCATED AT 250 WORDS)
Targeting of pancreatic islets of severe combined immunodeficient mice by passive transfer of allogeneic spleen cells from non-obese diabetic mice.
The precise role of immune cells in beta cell killing and their manner of invasion of pancreatic islets in insulin-dependent diabetes mellitus (IDDM) are unclear. We have attempted to target pancreatic islets of severe-combined immunodeficient (SCID) mice with spleen cells from diabetic and non-diabetic female non-obese diabetic (NOD) mice given i.p. or i.v. Pancreatic, liver and kidney sections of SCID mice were assessed histologically for the presence of donor cells. The presence of raised levels of serum Ig was also used as an index of engraftment of donor cells in the periphery of SCID mice. All six SCID mice which received i.v. spleen cells from normal Swiss mice died within 2 weeks from graft versus host disease (GVHD) whereas five out of nine mice survived for 30 days after i.p. injection. No deaths were recorded after i.v. or i.p. injection of spleen cells from NOD mice. Pancreatic islets of four out of six SCID recipients of diabetic and three out of five recipients of non-diabetic spleen cells following i.p. injection showed lymphocytic infiltrates in the peri-islet and perivascular regions. All SCID mice which received i.v. spleen cells from diabetic (six SCID recipients) and non-diabetic NOD mice (seven SCID recipients) showed peri-islet and perivascular infiltrates in their pancreas. Immunohistochemical analysis showed that the islet engrafted cells were of CD4 and CD8 phenotype. Donor cells were also observed in the exocrine pancreas of some recipients. A majority of mice showed various degrees of lymphocytic aggregates in the perivascular regions of the liver but not in the kidney.(ABSTRACT TRUNCATED AT 250 WORDS)
Cytoskeletal alterations in human fetal astrocytes induced by interleukin-1 beta.
Previous studies in this and other laboratories have shown that interleukin-1 beta (IL-1 beta) is a selective and potent activator of human astrocytes with respect to induction of cytokines and hematopoietic growth factors. To study the effect of recombinant human IL-1 beta (rhIL-1 beta) on astrocyte morphology, glial fibrillary acidic protein (GFAP) and vimentin expression, and actin organization, we conducted a systematic survey using dissociated human fetal astrocyte cultures. Within hours of stimulation with IL-1 beta, the majority of astrocytes converted from flat, polygonal cells to small, contracted, highly branched cells. This change in morphology was more striking when serum was eliminated from the medium. Complete dissolution of filamentous actin occurred simultaneously with the change in cell shape, as demonstrated by fluorescein-phalloidin binding. These "activated" astrocytes displayed intense GFAP and vimentin immunoreactivity in the small perikarya and processes. In contrast, the large, flat astrocytes in control cultures showed diffuse pale immunoreactivity for GFAP and vimentin. To quantify the changes in GFAP and vimentin content with IL-1 beta stimulation, densitometric analyses of northern and western blots were performed. Northern blot analysis of IL-1 beta-stimulated astrocytes revealed a transient, marked decrease in steady-state levels of mRNA for GFAP, vimentin, and microtubule-associated protein 4. The decrease in mRNA levels was evident by 4-8 h and fell to the lowest level at 16-24 h (80-98% decrease by densitometry) with partial recovery by 72 h. By immunoblotting, a significant decrease in both GFAP and vimentin protein content was observed after IL-1 beta stimulation. Furthermore, metabolic labeling studies revealed an almost total loss of GFAP synthesis following stimulation with IL-1 beta for 16 h. These observations are consistent with the idea that increases in immunoreactivity were related to factors such as redistribution of epitope, rather than increases in total protein content. We hypothesize that in IL-1 beta-stimulated astrocytes, synthesis of other proteins, e.g., inflammatory cytokines, occurs at the expense of structural proteins and that the decrease in content of cytoskeletal proteins may reflect an "activated" state of astrocytes.
Molecular characterization of a novel human gene, SEC13R, related to the yeast secretory pathway gene SEC13, and mapping to a conserved linkage group on human chromosome 3p24-p25 and mouse chromosome 6.
We previously described sequence tags from 58 novel directionally cloned human cDNAs from an enriched retinal pigment epithelial cell line library (Gieser and Swaroop, 1992). The nucleotide sequence of one of the cDNA clones, AA35 (D3S1231E), showed strong homology to the yeast SEC13 gene, required for vesicle biogenesis from endoplasmic reticulum during the transport of proteins. We have designated the human gene SEC13R (SEC13-Related). The amino acid sequence of the SEC13R gene product shows 70% similarity to yeast Sec13p, suggesting that SEC13R may be the human homolog of SEC13. The deduced polypeptide sequence contains several beta-transducin like 'WD40' repeats, and is rich in serine and threonine residues. The 1.4 kb transcript of SEC13R is detected by Northern analysis in many human tissues. However, RT-PCR analysis using two primer sets from different regions of the gene suggests differential expression of alternately spliced transcripts in various tissues. Somatic cell hybrid and in situ hybridization studies localized the SEC13R gene to human chromosome 3p24-p25. A related sequence was mapped to chromosome 18q11.2-q12. SEC13R was physically mapped to a yeast artificial chromosome (YAC) clone spanning the D3S720 marker from the region of the Von Hippel-Lindau disease locus. The mouse Sec13r gene was mapped to the conserved linkage group on chromosome 6 that corresponds to human chromosome 3p24-p25.
Conjugated linoleic acid (9,11- and 10,12-octadecadienoic acid) is produced in conventional but not germ-free rats fed linoleic acid.
Conjugated linoleic acid (CLA) is an anticarcinogen in several model animal systems. Conjugated linoleic acid occurs naturally in food and is present at higher concentrations in products from ruminant animals. Given that certain rumen microorganisms produce CLA from free linoleic acid, we studied the effect of feeding free or esterified linoleic acid on tissue CLA concentrations using conventional and germ-free rats. Conventional rats were fed a 5% (wt/wt) corn oil control diet alone or supplemented with 5% free linoleic acid or 8.63% corn oil (equivalent to 5% linoleic acid in triglyceride). Germ-free rats were fed autoclavable nonpurified diet alone or supplemented with 5% free linoleic acid. Analyses of CLA concentrations were performed on lipids extracted from liver, lung, kidney, skeletal muscle and abdominal adipose tissue, and on liver phospholipid and neutral lipid fractions. Tissue CLA concentrations were higher in conventional rats fed free linoleic acid (the major isomers were cis-9, trans-11 and trans-9, cis-11) than in control animals. Conjugated linoleic acid concentrations in free linoleic acid-fed rats were maximal at 4 wk, and levels were 5-10 times higher than those of controls. Elevated CLA concentrations were also observed in liver phospholipid and neutral lipid fractions. In contrast, CLA concentrations in the tissues of germ-free rats were not affected by diet. Feeding the corn oil-fortified diet to conventional rats did not increase CLA concentration in the tissues.(ABSTRACT TRUNCATED AT 250 WORDS)
Theoretical complex Stark energies of hydrogen by a complex-scaling plus B-spline approach.
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Expression of cytosolic retinoid-binding protein genes in human skin biopsies and cultured keratinocytes and fibroblasts.
Using reverse transcription coupled to polymerase chain reaction we have studied the mRNA expression of serum retinol-binding protein and cytosolic receptors for retinol and retinoic acid in skin biopsies, and in cultured epidermal keratinocytes and dermal fibroblasts. Transcripts for cellular retinol-binding protein (CRBP) I and cellular retinoic-acid-binding protein (CRABP) I were found in normal skin, keratinocytes, and fibroblasts. CRABP II transcripts were detected in skin and keratinocytes. A decreased mRNA expression of CRABP I and an increased mRNA expression of CRABP II were found in lesional psoriatic skin compared with uninvolved skin. mRNA transcripts for serum retinol-binding protein (s-RBP) were detected in all tissues and cells. The biological importance of s-RBP expression in keratinocytes and fibroblasts is not known, but hypothetically this protein may be involved in the intracellular shuttling of retinol and retinoic acid, or in the retransportation of cellular retinoids into the extracellular space.
Induction of chondrogenesis: requirement for synergistic interaction of basic fibroblast growth factor and transforming growth factor-beta.
Interactions between the epithelial anlage of the developing mouse inner ear and its associated periotic mesenchyme control the differentiation of the cartilaginous otic capsule. Transforming growth factor-beta 1 (TGF-beta 1) is a naturally occurring signal peptide that is present in these tissues at times of active differentiation and morphogenesis. Previous studies have shown that TGF-beta 1 alone is not a sufficient stimulus to initiate chondrogenesis in cultured periotic mesenchyme. In this study, we provide evidence that basic fibroblast growth factor (bFGF) can elicit a specific but limited chondrogenic response in cultured periotic mesenchymal cells. We also demonstrate that simultaneous addition of bFGF and TGF-beta 1 to cultured periotic mesenchyme results in a full chondrogenic response comparable to that which occurs when periotic mesenchyme is grown in the presence of its natural inductor tissue (i.e. otic epithelium). Utilizing antibodies directed against bFGF, we show localization of endogenous bFGF in the otic epithelium in vivo and in mixed epithelial-mesenchymal cultures. Additionally, we demonstrate the presence of FGF-like activity in medium conditioned by otic epithelium. Blocking of epithelial elicited chondrogenesis by a combination of both alpha bFGF and alpha TGF-beta 1 antibodies provides further evidence of the necessity for these growth factors in the chondrogenic differentiation of periotic mesenchyme in vitro. Our results suggest a role for both bFGF and TGF-beta 1 in the regulation of chondrogenesis during otic capsule formation in situ.
Pericardiocentesis and drainage by a silicon rubber line without echocardiographic guidance. Experience in 55 consecutive patients.
Pericardiocentesis and drainage with a silicon rubber line were performed safely in 55 patients via the subxiphoid (80 times) or apical (1 time) approach at the bedside without echocardiographic guidance. The subxiphoid approach, which is close to the posterior of the sternum or the costal cartilage with a small acute-angle (15-30 degrees) to the parietal pericardium in front of the right ventricle, can avoid injuring the heart, liver and lungs. A line with sideholes near the distal end, which provides convenient drainage and has better histocompatibility, can be kept in place for a long period of time (1-82 days). It was also convenient to inject air, drugs or fluid through the line. There were no procedure-related serious complications or deaths. Emergency drainage with the Seldinger technique, especially for small to moderate effusions, is safe and simple.
[Detection of B-cell clonality in paraffin-embedded bone marrow biopsy specimens by polymerase chain reaction].
In order to estimate the clonality of B-cell lymphoproliferative disorders, polymerase chain reaction (PCR) was used to detect the Ig heavy chain gene rearrangement in DNA extracted from formalin-fixed and paraffin-embedded bone marrow biopsies. 16 out of 19 cases (84%) of B-CLL (n = 9), multiple myeloma (n = 7) and B-non-Hodgkin's lymphoma (n = 3) showed sharp monoclonal bands, while polyclonal smears or no PCR-products were observed in non-neoplastic bone marrow or AML specimens. The results showed that the DNA from paraffin-embedded bone marrow biopsies could be used to detect monoclonal IgH rearrangement by PCR method. This forms the basis for the studies of minimal residual disease or B-ML bone marrow infiltrated diseases.
Studies on reducing and reinforcing manipulation of acupuncture during the past 4 years.
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[Clinical application of YAG laser in nasal endoscopic surgery].
In this study, we assessed the value of the clinical application of YAG laser in 32 cases of nasal endoscopic surgery. It was found that the operation performed under the nasal endoscope with YAG laser has several advantages including less bleeding, clear observation, simplified performance, thorough excision and less postoperative pain. But it also has some disadvantages, such as laser-induced lesion in the surrounding normal tissues, easy pollution and damage of the opto-guide fiber.
[Immunological detection of organophosphate resistance of Culex mosquitoes using anti-esterase monoclonal antibody].
Different strains of Culex mosquitoes (Cx. pipiens quinquefasciatus and Cx. Pipiens pallens) were immunologically-detected for organophosphate resistance with anti-esterase monoclonal antibody, and the results were compared with those detected by bioassay and biochemical microplate assay. It was found that the resistance-detection-rate detected by immunologic methods were higher than the corresponding data detected by the biochemical method, and the levels of resistance detected by sandwich-ELISA were higher than the corresponding levels detected by bioassay and the biochemical method. The thresholds for resistance in sandwich-ELISA were (at absorbence 450) > or = 0.5, and in microplate assay were (at absorbence 590) > or = 2.5 x 10(-3) mumol/min.mg protein. Dot-ELISA method was developed to meet the requirement of the field test and proved to be fast and convenient, especially in the detection of samples with higher esterase activity.
Comparative observations on the types of atrophic gastritis and biopsy pathology of gastric mucosa.
300 cases of chronic atrophic gastritis were classified into 3 types of qi-deficiency of spleen and stomach, yang-deficiency of spleen and stomach, and yin-deficiency of spleen and stomach according to syndrome differentiation of traditional Chinese medicine. The biopsy specimens of gastric mucosa were examined histopathologically and for the presence of pyloric spirobacteria. The results showed that the type of yin-deficiency was the most serious in the degree of pathologic changes, or the glandular atrophy and the interstitial inflammatory cellular infiltration, while the rate of pyloric spirobacteria presence was comparable in all types.
Studies on reducing and reinforcing manipulation of acupuncture during the past 4 years (continued).
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[Clinicopathologic and immunohistochemical studies of midline malignant reticulosis in palate].
Fourteen patients (10 males and 4 females) with midline malignant reticulosis (MMR) in palate were studied. The symptoms were most commonly swelling and ulceration. In some cases, the lesion involved nasal, maxillary sinus and facial skin, but no regional lymph nodes were involved. A necrotizing infiltration of atypical lymphoid cells (ALCs) mixed with inflammatory cells was confirmed as the histopathologic feature of MMR. In 2 of 14 cases, ALCs formed a continuous sheet in several high-power fields with a few inflammatory cells. This overlapping feature may be a transitional form between MMR and malignant lymphoma. Immunohistochemical staining results indicated that in 10 of 14 cases, ALCs were CD3 positive; in 7 of 14 cases, ALCs were UCHL1 positive; and in all 14 cases, ALCs were L26 negative. We deem MMR a T-cell lymphoma in substance, but it should be treated as a histopathologic entity distinct from malignant lymphoma.