Biomedical subjects

H Ma

Publications and source records attributed to H Ma.

At least 145 records · Page 8Linked to original sources

PubMed1999

Lp82 is the dominant form of calpain in young mouse lens.

The purpose of this study was to characterize Lp82 calpain in normal mouse. Lp82 is a lens-specific, calcium-activated isozyme from the calpain super family of cysteine proteases (EC 34.22.17). RT-PCR and molecular cloning were performed on total RNA from 12 day-old mice. Lp82 and m-calpain protein levels and proteolytic activities in lenses were measured by casein zymography, immunoblotting, and ELISA after partial purification by DEAE-HPLC. The 2334-bp cDNA encoding for mouse Lp82 contained a single large open reading frame encoding a protein of 709 amino acid residues with a calculated molecular weight of 82.2 kDa and a predicted pI of 5.8. The amino acid sequence of mouse lens Lp82 was 99% homologous to rat lens Lp82. As in rat, mouse lens Lp82 showed a unique N -terminus and deletion of the IS1 and IS2 regions. In contrast to rat, Lp82 was the dominant calpain in young mouse lens. Lp82 was lens-specific, and the lens nucleus contained the highest specific activity of Lp82 and very little m-calpain. Endogenous Lp82 in lens soluble proteins was activated by addition of calcium and caused limited proteolysis of crystallins even in the presence of large amounts of recombinant domain I from the natural calpain inhibitor calpastatin. Loss of Lp82 protein accompanied aging of mouse lens. Lp82 may be responsible for a major portion of crystallin proteolysis occurring during normal lens development and maturation, or during cataract formation in young mice.

Amino Acid Sequence↗

PubMed1999

Decreased sensitivity of lens-specific calpain Lp82 to calpastatin inhibitor.

The purpose of the present investigation was to test three calpain inhibitors (recombinant calpastatin domain I, E64, and SJA6017) against Lp82 calpain in rat lenses. Lp82 is a lens-specific isoenzyme from the calpain super family of calcium-activated, cysteine proteases (EC 34.22.17). Lp82 and m-calpain proteolytic activities and protein levels were measured by casein zymography and immunoblotting. Activity of endogenous Lp82 against vimentin was also tested by in vitro incubation of rat lens soluble and insoluble fractions with calcium. Most of the Lp82 activity could be inhibited by irreversible inhibitor E64 and reversible inhibitor SJA6017. However, a major finding of the present investigation was that Lp82 in the soluble and insoluble fractions of the lens was less sensitive to inhibition by recombinant domain I from the endogenous tissue inhibitor of ubiquitous calpains, calpastatin, than m-calpain. By using recombinant calpastatin to inhibit endogenous lens m-calpain, we were able to demonstrate the first example of a substrate for Lp82, vimentin. These data suggest that Lp82-induced proteolysis in rodent lenses may occur even in the presence of calpastatin.

Animals↗

PubMed1999

Electroactive hemoglobin-surfactant-polymer biomembrane-like films.

Polyions complex 2C12N+ PSS- was prepared by reacting poly(sodium styrenesulfonate) (Na+ PSS-) with didodecyldimethylammonium bromide (2Cl2N+ Br-). Stable thin films made from 2C12N+ PSS- with incorporated redox protein hemoglobin (Hb) on pyrolytic graphite (PG) electrodes were then characterized by electrochemistry and other techniques. Cyclic voltammetry (CV) of Hb-2C12N+ PSS- films showed a pair of well-defined and nearly reversible peaks for HbFe(III)/Fe(II) couple at about -0.17 V vs. saturated calomel electrode (SCE) in pH 5.5 buffers. The electron transfer rate between Hb and PG electrode was greatly facilitated in microenvironment of 2C12N+ PSS- films. Positions of Soret absorption band suggest that Hb keeps its secondary structure similar to its native state in 2C12N+ PSS- films at the medium pH. The results of X-ray diffraction and differential scanning calorimetry (DSC) suggest synthesized lipid 2C12N+ PSS- films have an ordered bilayer structure intercalated between PSS- polyion layers, and the incorporated Hb expands the layer spacing of the films. HbFe(I), a highly reduced form of Hb, might also be produced in these films at about -1.09 V, and could be used to catalytically reduce organohalide pollutants.

Calorimetry, Differential Scanning↗

PubMed1999

Mutations within the Autographa californica nucleopolyhedrovirus FP25K gene decrease the accumulation of ODV-E66 and alter its intranuclear transport.

Previous reports indicate that mutations within the Autographa californica nucleopolyhedrosis virus FP25K gene (open reading frame 61) significantly reduce incorporation of enveloped nucleocapsids into viral occlusions. We report that FP25K is a nucleocapsid protein of both the budded virus (BV) and occluded virus (ODV), and we describe the effects of two FP25K mutations (480-1 [N-terminal truncation] and FP-betagal [C-terminal fusion]) on the expression and cellular localization of ODV-E66 and ODV-E25. Significantly decreased amounts of ODV-E66 are detected in cells infected with 480-1 or FP-betagal viral mutants, even though during FP-betagal infection, steady-state levels of ODV-E66 transcripts remain unchanged. While ODV-E66 is normally detected in intranuclear microvesicles and ODV envelopes by 24 h postinfection (p.i.), ODV-E66 remains cytosolic throughout infection in cells infected with 480-1 virus (up to 96 h p.i.), and its intranuclear localization is not detected until 96 h p.i. in cells infected with the FP-betagal mutant virus. The nuclear localization of ODV-E25 is not affected during infection by the FP-betagal mutant; however, its trafficking is significantly delayed during infection by the 480-1 mutant. Temporal Western blot analyses of cell lysates show that both 480-1 and FP-betagal mutant virus infections result in altered accumulation patterns of several structural proteins, including gp67, BV/ODV-E26, and the major capsid protein p39. In addition to BV/ODV-E26, ODV-E66 and gp67 may interact with FP25K, and ODV-E25 and p39 may also be components of a protein complex containing ODV-E66 and FP25K. Together, these data suggest that FP25K and its associated protein complex(es) may play an important role in the targeting and intracellular transport of viral proteins during infection.

Amino Acid Sequence↗

PubMed1999

Multiple signal input and output domains of the 160-kilodalton nuclear receptor coactivator proteins.

Members of the 160-kDa nuclear receptor coactivator family (p160 coactivators) bind to the conserved AF-2 activation function found in the hormone binding domains of nuclear receptors (NR) and are potent transcriptional coactivators for NRs. Here we report that the C-terminal region of p160 coactivators glucocorticoid receptor interacting protein 1 (GRIP1), steroid receptor coactivator 1 (SRC-1a), and SRC-1e binds the N-terminal AF-1 activation function of the androgen receptor (AR), and p160 coactivators can thereby enhance transcriptional activation by AR. While they all interact efficiently with AR AF-1, these same coactivators have vastly different binding strengths with and coactivator effects on AR AF-2. p160 activation domain AD1, which binds secondary coactivators CREB binding protein (CBP) and p300, was previously implicated as the principal domain for transmitting the activating signal to the transcription machinery. We identified a new highly conserved motif in the AD1 region which is important for CBP/p300 binding. Deletion of AD1 only partially reduced p160 coactivator function, due to signaling through AD2, another activation domain located at the C-terminal end of p160 coactivators. C-terminal coactivator fragments lacking AD1 but containing AD2 and the AR AF-1 binding site served as efficient coactivators for full-length AR and AR AF-1. The two signal input domains (one that binds NR AF-2 domains and one that binds AF-1 domains of some but not all NRs) and the two signal output domains (AD1 and AD2) of p160 coactivators played different relative roles for two different NRs: AR and thyroid hormone receptor.

Animals↗

PubMed1999

Comparison of SERCA1 and SERCA2a expressed in COS-1 cells and cardiac myocytes.

Cultured COS-1 cells, as well as chicken embryonic and neonatal rat cardiac myocytes, were infected with recombinant adenovirus vectors to define limiting factors in the expression and Ca2+ transport function of recombinant sarcoplasmic-endoplasmic reticulum Ca(2+) (SERCA) isoforms. Titration experiments showed that all COS-1 cells and myocytes in culture could be infected by an adenovirus titer of 10 plaque-forming units (pfu) per seeded cell. Raising the adenovirus titer further yielded higher protein expression up to an asymptotic limit for functional, membrane-bound SERCA protein. The asymptotic behavior of SERCA expression was not transcription related but was due to posttranscriptional events. The minimal (-268) cardiac troponin T (cTnT) promoter was a convenient size for adenovirus vector construction and manifested tight muscle specificity. However, its efficiency was lower than that of the nonspecific cytomegalovirus (CMV) promoter. At any rate, identical maximal levels of SERCA expression were obtained with the CMV and the cTnT promoter, as long as the viral titer was adjusted to compensate for transcription efficiency. A maximal threefold increase of total SERCA protein expression over the level of the endogenous SERCA of control myocytes was reached (a sevenfold increase compared with the endogenous SERCA of the same infected myocytes due to reduction of endogenous SERCA after infection). In contrast with previous reports [Ji et al. Am. J. Physiol. 276 (Heart Circ. Physiol. 45): H89-H97, 1999], a higher kinetic turnover was demonstrated for the SERCA1 compared with the SERCA2a isoform as shown by a 5.0- versus 2.6-fold increase in calcium uptake rate accompanying maximal expression of recombinant SERCA1 or SERCA2a, respectively. This information is deemed necessary for studies attempting to modify myocardial cell function by manipulation of SERCA expression.

Adenoviridae↗

PubMed1999

Protective effect of quinaprilat, an active metabolite of quinapril, on Ca2+-overload induced by lysophosphatidylcholine in isolated rat cardiomyocytes.

We examined the effects of quinaprilat, an active metabolite of quinapril (an angiotensin converting enzyme (ACE) inhibitor) on the increase in intracellular concentration of Ca2+ ([Ca2+]i) (Ca2+-overload) induced by lysophosphatidylcholine (LPC) in isolated rat cardiomyocytes. LPC (15 microM) produced Ca2+-overload with a change in cell-shape from rod to round. Quinaprilat but not quinapril at 20 or 50 microM attenuated the LPC-induced increase in [Ca2+]i and the change in cell-shape in a concentration-dependent manner. Since quinaprilat has an inhibitory action on ACE and quinapril has practically no inhibitory action on ACE, it is likely that the inhibitory action of quinaprilat on ACE is necessary for the protective effect of the drug against LPC-induced changes. We therefore examined the effects of enalapril (another ACE inhibitor with the weak inhibitory action on ACE) and enalaprilat (an active metabolite of enalapril with an inhibitory action on ACE) on the LPC-induced changes. Both enalapril and enalaprilat attenuated the LPC-induced Ca2+-overload, suggesting that the inhibitory action on ACE may not mainly contribute to the protective effect of ACE inhibitors against LPC-induced Ca2+-overload. This suggestion was supported by the fact that neither ACE (0.2 U/ml) nor angiotensin II (0.1-100 microM) increased [Ca2+]i in isolated cardiomyocytes. Furthermore, application of bradykinin (0.01-10 microM) did not enhance the protective effect of quinaprilat against LPC-induced changes. LPC also increased release of creatine kinase (CK) from the myocyte markedly, and quinaprilat but not quinapril attenuated the LPC-induced CK release. Unexpectedly, both enalapril and enalaprilat did not attenuate the LPC-induced CK release. Neither quinapril nor quinaprilat changed the critical micelle concentration of LPC, suggesting that these drugs do not directly bind to LPC. We conclude that quinaprilat attenuates the LPC-induced increase in [Ca2+]i, and that the protective effect of quinaprilat on the LPC-induced change may not be related to a decrease in angiotensin II production or an increase in bradykinin production.

Angiotensin-Converting Enzyme Inhibitors↗

PubMed1999

[Study of NAIP gene in spinal muscular atrophy].

OBJECTIVE: Investigating the correlation between the genotype and spinal muscular atrophy(SMA) clinical phenotype. METHODS: Neuronal apoptosis inhibitory protein(NAIP) gene analysis was performed by PCR amplification of exon 5 in 13 unrelated SMA patients with deletions of SMN gene. RESULTS: Two Type-I patients(2 M5, 40%) lacked the NAIP exon 5. CONCLUSION: The results suggest a possible correlation between NAIP gene deletion and the disease severity of SMA.

Child↗

PubMed1999

[Morphological study of CD3+ dendritic epidermal T cells in human skin and scar].

OBJECTIVE: To investigate the CD3/TCR-bearing T cell in the epidermis and explore the influence of the dendritic epidermal T cells on regeneration and differentiation of epidermal cells. METHODS: The CD3/TCR-bearing T cells in epidermis of human skin and cicatrix were examined by means of histomorphology and immunohistochemistry. RESULTS: It was found that the density of CD3/TCR-bearing T cells in human skin and scar tissue had great dissimilarity. CD3+ dendritic epidermal T cells increased in hyperplastic scar. Such result was not observed in atrophic scar. CONCLUSION: Our findings indicate CD3+ dendritic epidermal T cells regulate the generation and differentiation of epidermal cells. CD3+ dendritic epidermal T cells may play an important role in the maintenance of the morphology of epidermal tissue.

Adult↗

PubMed1999

[Cleft lip repair using a lateral columellar flap].

OBJECTIVE: Since the important cosmetic structures of a good looking lip, viz. the philtrum column, philtrum dimple and Cupid's bow of a cleft lip repaired with the commonly used methods, are still defective in various degrees, a new method is designed. METHODS: On the medial side of the cleft, the short philtrum is lengthened and the upwardly displaced Cupid's bow maintained in a lowered position by a rectangular flap transplanted from the lateral surface of the columella. No lip flap has been formed on the lateral side of the cleft. RESULTS: In a period of three years (1996-1998), 23 cases of unilateral cleft lip, 9 cases of bilateral cleft lip and 14 cases of secondary deformities of cleft lip have been repaired with this new method. All of the repaired lips have normal looking philtrum column (limited by an uninterrupted straight-line scar), philtrum dimple and Cupid's bow. CONCLUSION: The lateral columellar flap method is a more rational surgical procedure for cleft lip repair in that it repairs tissue defect by transplanting tissue from the unexposed area. The appearance of the repaired lip is better than that of the commonly used methods.

Cleft Lip↗

PubMed1999

[A study on guided tissue regeneration and tissue engineering-transplantation of collagen membrane seeded with cultured hyaline chondrocytes].

OBJECTIVE: Lack of transplant material is a main problem in the repair and reconstruction of tissue or organ defects. To seek the succedaneum of transplants, we studied and used tissue-engineered growth hyaline cartilage. METHODS: The articular hyaline cartilage obtained from newborn calf within 6 hours of sacrifice was enzymatically dissolved. The chondrocytes were then seeded onto the medical collagen membrane of guided tissue regeneration in a 24 well plate in RPMI1640 medium. One week later, the chondrocyte medical collagen membrane of guided tissue regeneration complex was implanted subcutaneously into the back of nude mice. Eight weeks after implantation, the animals were sacrificed. RESULTS: The present study seems to demonstrate that the cartilage-like tissue was strong enough to be transferred after being implanted for 8 weeks. The cartilage-like tissue was proved to be tissue-engineered hyaline cartilage by HE stain. The chondrocytes could secrete chondroitin sulfate as proved by Lev-Spicer stain. CONCLUSION: Tissue engineering technique was used to make tissue-engineered cartilage in vitro. It will provide a new transplant material for the repair of tissue or organ defects.

Animals↗

PubMed1999

Congenital choledochal cyst with pancreatitis.

OBJECTIVE: To understand the relationship among congenital choledochal cyst, anomalous junction of pancreaticobiliary duct (AJPBD) and pancreatitis. METHODS: 25 children with choledochal cyst treated in our hospital were included in this study. Their ages ranged from 2 months to 14 years. Intraoperative cholangiography was performed in 24 children. Pancreatic samples obtained from the head and body of the pancreas at operation were observed under light and electron microscope. Serum amylase was analyzed one week before and after operation respectively. Bile amylase in the cyst and gallbladder was measured at operation. RESULTS: The incidence of AJPBD was 70%. Inflammatory cell infiltration and fibrous hyperplasia were seen in the pancreatic specimens of 5 children, of whom 4 had AJPBD. No ultrastructural changes were found in 2 children without AJPBD, while different changes were seen in 6 with AJPBD, including those showing no pathological changes under light microscope. All the changes became more severe as the age of the patient increased. The bile and serum amylase levels were higher in children with AJPBD than those without AJPBD (P < 0.05). CONCLUSION: Congenital choledochal cyst has a close relationship with AJPBD and pancreatitis. Pancreatic pathological changes have a long and chronic course from ultrastructural changes to macroscopic changes for the developing of pancreatitis.

Adolescent↗

PubMed1999

[Analysis of bone mineral density and relevant factors in 781 healthy people aged 15 to 50].

OBJECTIVE: To determine further the age to form peak bone mass (PBM) and the factor influencing it and bone mineral density (BMD) and to find an effective way to prevent from osteoporosis in the elderly. METHODS: BMD was measured in 781 people aged 15 to 50 years and relevant factors in lifestyle were investigated. Multivariate regression was used to analyze the relationship between BMD and lifestyles and to identify both risk and benefit factors for osteoporosis. RESULTS: For every ten-kilogram increase in body weight, BMD at varied sites increased by an amount of 0.028 - 0.056 g/cm(2) for males and 0.050 - 0.086 g/cm(2) for females, respectively. During their adolescence for males, for every increase of one grade in physical exercise, BMD at varied sites increased by an amount of 0.034 - 0.078 g/cm(2) in males and BMD at lumbar vertebra in males aged 15 - 24 correlated negatively with their total amount of alcohol drinking, and that at lumbar vertebra and of the whole body increased by about 0.035 g/cm(2) in females, which correlated negatively with their total amount of cigarette smoking. Female BMD at lumbar vertebra correlated reversely with their age of menarche and correlated positively with their average time of breast feeding. CONCLUSION: Increase in physical exercise during puberty could increase PBM for the adolescents and keeping adequate body weight and reducing cigarette smoking and alcohol drinking would benefit for the prevention from age-related osteoporosis.

Absorptiometry, Photon↗

PubMed1999

[Changes of CGRP activity and distribution in facial nucleus after facial nerve injury of guinea pig].

OBJECTIVE: In order to study the changes of CGRP activity in facial nucleus of guinea pig after facial nerve injury. METHOD: The activity of CGRP was detected by the technique of immunohistochemical and image analysis methods were used. RESULT: It was showed the there were CGRP-immunoreactivity (IR) in normal facial nucleus and its' subnucleus. An initial increase of CGRP-IR was noted and the content of CGRP under image analysis were significantly different when compared with control group (P < 0.01) at day 1 after injury of facial nerve. CGRP continued to rise to a maximal level at day 7 (P < 0.001). And after which it gradually decreased. We noticed distinct bundle-shaped CGRP fibers elongated to the peripheral part from ventral facial nucleus in the period of 14 d-35 d. CONCLUSION: It suggests that during the course of facial nerve regeneration following injury, CGRP changed characteristically. CGRP may play the regulational role in facial nerve regeneration.

Animals↗

PubMed1998

Spatial and temporal dynamics of DNA replication sites in mammalian cells.

Fluorescence microscopic analysis of newly replicated DNA has revealed discrete granular sites of replication (RS). The average size and number of replication sites from early to mid S-phase suggest that each RS contains numerous replicons clustered together. We are using fluorescence laser scanning confocal microscopy in conjunction with multidimensional image analysis to gain more precise information about RS and their spatial-temporal dynamics. Using a newly improved imaging segmentation program, we report an average of approximately 1,100 RS after a 5-min pulse labeling of 3T3 mouse fibroblast cells in early S-phase. Pulse-chase-pulse double labeling experiments reveal that RS take approximately 45 min to complete replication. Appropriate calculations suggest that each RS contains an average of 1 mbp of DNA or approximately 6 average-sized replicons. Double pulse-double chase experiments demonstrate that the DNA sequences replicated at individual RS are precisely maintained temporally and spatially as the cell progresses through the cell cycle and into subsequent generations. By labeling replicated DNA at the G1/S borders for two consecutive cell generations, we show that the DNA synthesized at early S-phase is replicated at the same time and sites in the next round of replication.

3T3 Cells↗

PubMed1998

Estrogen-induced production of a peroxisome proliferator-activated receptor (PPAR) ligand in a PPARgamma-expressing tissue.

Peroxisome proliferation has been associated with carcinogenesis in the liver, and estrogen intake has been associated with increased risk of cancer in the hormone target tissues. Estrogen-induced peroxisome proliferation has been observed in an estrogen target tissue, the uropygial gland in the duck. To elucidate the molecular mechanism of this process, we previously isolated the cDNA of peroxisome proliferator-activated receptor gamma1 (PPARgamma1) from the duck uropygial gland and found that its expression was high exclusively in this tissue of duck. However, the nature of the ligand for PPARgamma1 and how estrogen might enhance PPARgamma1-regulated gene expression were not known. Here we demonstrate that estrogen treatment of animals enhanced the metabolism of arachidonic acid in the uropygial gland. Conversion of prostaglandin D2 to a metabolite was induced by estradiol treatment preceding peroxisome proliferation. High performance liquid chromatography and TLC analyses showed that the metabolite behaved chromatographically similar to prostaglandin J2 and Delta12-prostaglandin J2. Gas chromatography/mass spectrometry revealed a striking similarity of the metabolite to Delta12-prostaglandin J2, the only form among the J2 series whose natural occurrence has been detected. Furthermore, this metabolite was able to activate duck PPARgamma1 to the same extent as the same concentrations of Delta12-prostaglandin J2 and 15-deoxy-Delta12, 14-prostaglandin J2, whereas under the same conditions, prostaglandin D2 was not effective. The results suggest that estrogen treatment induced the formation of a prostaglandin D2 metabolite that activated duck PPARgamma1, causing the induction of peroxisome proliferation in the duck uropygial gland.

Animals↗

PubMed1998

Flowering time: from photoperiodism to florigen.

An Arabidopsis blue-light receptor, Cry2, has been found to play a critical role in the photoperiodic control of flowering time; and genes have been identified that may control the production of a transmissible flower-inducing signal, which may turn out to be the long-elusive putative flowering hormone 'florigen'.

Arabidopsis↗

PubMed1998

Peroxisome proliferator-activated receptor gamma1 (PPAR-gamma1) as a major PPAR in a tissue in which estrogen induces peroxisome proliferation.

Estradiol administration induces peroxisome proliferation and the production of 3-hydroxy fatty acid pheromones in the uropygial glands of the duck, but not in the goose gland, which does not produce such pheromones. We isolated a peroxisome proliferator-activated receptor (PPAR)gamma1 cDNA from a duck uropygial gland cDNA library. Northern blots revealed two transcripts, PPAR gamma1 and gamma2, and showed that PPAR gamma was expressed at higher levels than PPAR alpha in the uropygial gland of the duck. Although PPAR gamma2 was expressed in both duck and goose uropygial gland, PPAR gamma1 was expressed only in the duck gland, which responds to estrogen by peroxisome proliferation. In NIH 3T3 transfected cells, PPAR gamma1 was activated by peroxisome proliferators such as Wy-14643, clofibric acid and Ly-171883 causing induction of the target marker gene. By cotransfection with a plasmid containing alpha-cis-retinoic acid receptor RXR alpha, the induction increased up to 9-fold. These results suggest that PPAR gamma1 may be involved in peroxisome proliferation while PPAR gamma2 may be involved in lipid metabolism.

3T3 Cells↗