Search PubMed⌕ Search

Biomedical subjects

C C Hsu

Publications and source records attributed to C C Hsu.

At least 55 records · Page 3Linked to original sources
PubMed2000

A novel method for expression and large-scale production of human brain l-glutamate decarboxylase.

l-Glutamate decarboxylase (GAD; EC 4.1.1.15) is the rate-limiting enzyme involved in the synthesis of gamma-aminobutyric acid (GABA), the major inhibitory neurotransmitter in the mammalian brain. Imbalance in the conversion of glutamate to GABA has been implicated in a host of human diseases. Studies on the structure, function, and therapeutic use of GAD have been precluded by insufficient quantities of purified active enzyme. Here we report a novel methodology for the expression and large-scale production of enzymatically active, pure, recombinant human GAD65 and GAD67. This method circumvents the sequestering of expressed protein into insoluble inclusion bodies and reduces production of truncated proteins. The availability of sufficient quantities of purified HGAD65 and HGAD67 has allowed for the production of specific polyclonal antibodies that discriminate between the two isoforms. This methodology, in addition to providing key human brain enzymes, may be generally applicable to other systems.

Brain↗
PubMed2000

Hereditary haemorrhagic telangiectasia: hepatic lesions demonstrated with colour Doppler and power Doppler sonography.

Visceral vascular abnormalities in patients with hereditary haemorrhagic teleangiectasia (HHT) include teleangiectasis, arteriovenous fistula (AVF), and aneurysms. Sonographic findings and colour Doppler ultrasound (CDU) features of HHT in the liver have been described in a few reports. Presented here is the CDU finding of liver with HHT involvement in two patients, aged 52 and 29 years, respectively. Power Doppler sonography was also performed in the first patient in whom both CDU and power Doppler sonography could demonstrate the abnormal vascular channels in the liver. In the second patient, aneurysms in the left hepatic lobe and of the splenic artery were noted. Angiography was performed in both patients and confirmed the abnormal vascular lesions demonstrated by CDU and/or power Doppler sonographic findings.

Adult↗
PubMed2000

Agitation in nursing home residents: the role of gender and social context.

We investigated the relationship among gender of resident, staff social interaction, and agitation in 46 (31 male and 15 female) nursing home residents with clinically significant agitation. Direct observations were conducted of resident behaviors and environmental contextual events using a computer-assisted, real-time observational system. The system recorded frequency, duration, and temporal sequencing of events. Results show that female residents displayed almost three times the amount of agitation as male residents (35% vs. 13% of total observation time, respectively), although men in the study were more likely to receive psychoactive drugs for their agitation. Staff spent similar amounts of time verbally interacting and touching male and female residents. Sequential analyses were conducted to examine the likelihood of staff verbal and touch interactions both preceding and following resident agitation using Bakeman and Quera's (1995) SDIS-GSEQ program. Results suggest that staff touch and verbal interaction elicit agitation in a significant proportion of residents. Once agitation occurs, staff were likely to respond by interacting verbally, but not physically, with the resident.

Aged↗
PubMed2000

Epidemiological study of 134 subjects with allergic conjunctivitis.

At times it is difficult to distinguish among the subtypes of chronic allergic conjunctivitis. A prospective study with 134 patients was carried out. Patients were diagnosed with vernal keratoconjunctivitis (VKC), atopic keratoconjunctivitis (AKC) or perennial allergic conjunctivitis (PAC). Demographic information was also collected including, age at symptom onset, sex, history of non-ocular allergy, and history of family allergy. Forty-six percent were diagnosed with VKC, 40% with AKC, 8% with PAC, and 6% were not specifically diagnosed. This distribution information as well as the demographic information collected demonstrates that there are patterns within each subgroup of chronic ocular allergy sufferers. These findings will be helpful in the diagnosing and proper classification of ocular allergic conditions.

Brazil↗
PubMed2000

Menstruation and reproduction in women with polychlorinated biphenyl (PCB) poisoning: long-term follow-up interviews of the women from the Taiwan Yucheng cohort.

BACKGROUND: In 1979 in Taiwan, about 2000 people were poisoned by cooking oil contaminated with heat-degraded polychlorinated biphenyls (PCBs). Victims developed chloracne and other symptoms; the illness is called Yucheng (oil disease). The chemicals persist in the victims' bodies and some are toxic to the female reproductive system in experimental animals. METHODS: In 1993-1994, we identified the living Yucheng women and neighbourhood controls and interviewed them about their reproductive experience. Of the 596 living Yucheng women aged 30-59 years, we found 368, and interviewed 356. We identified 329 controls, and interviewed 312. RESULTS: Of the Yucheng women, 16% reported abnormal menstrual bleeding compared to 8% (P < 0.05) of control women; 4.2% versus 1.7% reported a stillbirth since 1979 (P = 0.068). Other characteristics of the menses, fertility, frequency of intercourse, and age at menopause appeared unaffected. More of the Yucheng women reported that one of their offspring had died during childhood (10.2% versus 6.1%, P < 0. 05), and that they had decided to limit childbearing because of health problems (7% versus 2%, P = 0.01). CONCLUSIONS: These findings suggest that high level PCB/polychlorinated dibenzofuran (PCDF) exposure has some effect on female endocrine and reproductive function.

Adult↗
PubMed2000

Three different turkey luteinizing hormone receptor (tLH-R) isoforms I: characterization of alternatively spliced tLH-R isoforms and their regulated expression in diverse tissues.

Using combinations of reverse transcription-polymerase chain reaction (RT-PCR) and 5'- and 3'-rapid amplification of cDNA ends, three different, alternatively spliced, partial turkey LH receptor (tLH-R) cDNA isoforms were characterized from ovarian mRNA. The first cDNA (tLH-R(intact)) showed 98% and 72-75% similarity with chicken and mammalian LH-R sequences, respectively. The second cloned cDNA isoform (tLH-R(insert)) contained an in-frame TGA stop codon within an 86-base pair insertion that was located in the extracellular domain of the seven-transmembrane region. The tLH-R(insert) isoform could encode a truncated soluble protein isoform that lacked the transmembrane region. The third cDNA isoform truncated the transmembrane region (tLH-R(trunc)) and was derived by the deletion of the last exon by incomplete splicing. Generation of multiple transcripts by alternative splicing was elucidated by partial characterization of tLH-R genomic sequences. The differentially regulated expression of the tLH-R mRNA isoforms in nongonadal tissues and ovarian stromal tissues during various reproductive stages was quantified and analyzed by Northern blot and/or RT-PCR. Alternatively spliced tLH-R isoforms were differentially expressed in a tissue-specific manner in most of the tissues examined. The steady-state levels of tLH-R mRNA isoforms were relatively high in the hypothalamus and optic nerve and relatively low in the cortex, pituitary, and cerebellum when compared to levels in ovarian follicles. In nongonadal reproductive tissues, the steady-state levels of tLH-R mRNA isoforms were relatively high in the uterus and infundibulum and relatively low in the isthmus, oviduct, and magnum. In addition, in the nongonadal peripheral tissues, the steady-state levels of tLH-R isoforms were relatively high in the thyroid gland and relatively low in the spleen, adrenal gland, kidney, skin, bursa, and muscle. The present study suggests that the alternative splicing of LH-R transcripts occurs in a tissue-specific manner and has been evolutionarily conserved (similar results were obtained in chicken and swine). These results raise fundamental questions as to the function of LH-R isoforms in nongonadal tissues.

Alternative Splicing↗
PubMed2000

Areca nut extract up-regulates prostaglandin production, cyclooxygenase-2 mRNA and protein expression of human oral keratinocytes.

There are about 600 million betel quid (BQ) chewers in the world. BQ chewing is associated with increased incidence of oral cancer and submucous fibrosis. In this study, areca nut (AN) extract (200-800 microg/ml) induced the prostaglandin E(2) (PGE(2)) production by 1. 4-3.4-fold and 6-keto-PGF(1 alpha) production by 1.1-1.7-fold of gingival keratinocytes (GK), respectively, following 24 h of exposure. Exposure of GK to AN extract (>400 microg/ml) led to cell retraction and intracellular vacuoles formation. At concentrations of 800 and 1200 microg/ml, AN extract induced cell death at 21-24 and 32-52% as detected by MTT assay and cellular lactate dehydrogenase release, respectively. Interestingly, AN-induced morphological changes of GK are reversible. GK can still proliferate following exposure to AN extract. Cytotoxicity of AN extract cannot be inhibited by indomethacin (1 microM) and aspirin (50 microM), indicating that prostaglandin (PG) production is not the major factor responsible for AN cytotoxicity. PGE(2) exhibited little effect on the growth of GK at concentrations ranging from 100-1000 pg/ml. Stimulating GK production of PGs by AN extract could be due to induction of cyclooxygenase-2 (COX-2) mRNA expression and protein production. These results suggest that AN ingredients are critical in the pathogenesis of oral submucous fibrosis and oral cancer via their stimulatory effects on the PGs, COX-2 production and associated tissue inflammatory responses. AN cytotoxicity to GK is not directly mediated by COX-2 stimulation and PG production.

6-Ketoprostaglandin F1 alpha↗
PubMed2000

Percutaneous epididymal sperm aspiration versus microsurgical epididymal sperm aspiration for irreparable obstructive azoospermia--experience with 100 cases.

PURPOSE: This study investigated the sperm retrieval success rates, fertilization rates, pregnancy rates, and complications of percutaneous epididymal sperm aspiration (PESA) and microsurgical epididymal sperm aspiration (MESA) in cases of irreparable obstructive azoospermia. METHODS: During a period of 36 months, 100 men with irreparable obstructive azoospermia underwent 109 cycles of sperm retrieval procedures and intracytoplasmic sperm injection (ICSI). We routinely performed PESA first in each retrieval cycle; MESA and/or testicular sperm extraction (TESE) were performed if PESA failed. The sperm retrieval success rates, mean fertilization rates, and pregnancy rates of PESA and MESA were evaluated. RESULTS: PESA was performed in all 109 retrieval cycles with a successful sperm retrieval rate of 61%. When PESA failed to retrieve a sufficient number of viable sperm, MESA was subsequently performed with a sperm retrieval rate of 93%. Three cases, which had failed retrieval with both the PESA and MESA procedures, received TESE successfully. The rates of fertilization and pregnancy were 56% and 39% in the 66 PESA-ICSI cycles, respectively, and 47% and 45% in the 40 MESA-ICSI cycles. No significant differences were found in fertilization rates or pregnancy rates among the various sperm retrieval methods and obstruction etiologies. The overall mean fertilization rate and pregnancy rate were 51% and 41%, respectively. CONCLUSION: Both PESA and MESA can be used successfully to obtain sufficient sperm for ICSI. PESA cannot replace MESA in some cases as some epididymal pathologies prevent its success. The results of this study indicate that PESA should be the treatment of choice for patients with ductal obstruction distal to the epididymis, owing to its higher initial success rate. In contrast, patients with irreparable epididymal obstruction might achieve better success rates with MESA.

Epididymis↗
PubMed1999

Hydrogen peroxide induces premature acrosome reaction in rat sperm and reduces their penetration of the zona pellucida.

Recent studies have demonstrated that mammalian sperm are capable of generating reactive oxygen species (ROS) and that this activity is significantly accelerated in subfertile subjects. The observed decrease in penetration of zona-intact oocyte might be explained by chemical-induced ROS-related early onset of capacitation and premature acrosome reaction, but the mechanism is not clear. We determine whether zona-intact oocyte penetration capability in rat epididymal sperm was affected by premature acrosome reaction in rat sperm treated with hydrogen peroxide (H2O2) and calcium ionophore A23187 or H2O2 and lysophosphatidyl choline. Chlortetracycline fluorescence assay was used to study the status of acrosome reaction on epididymal sperm. The sperm-oocyte binding and penetration assay was used to evaluate the capability for zona pellucida penetration. There was a positive linear correlation between the frequency of acrosome-reacted sperm and capability of sperm-oocyte binding and penetration in zona-free oocytes. In the zona-intact oocytes, the sperm-oocyte penetration rate was suppressed as the proportions of acrosome-reacted sperm increased. In summary, this study showed that premature acrosome reaction reduced rat sperm's capability of penetrating zona-intact oocytes. However, this reduction is not seen in zona-free oocytes. These findings may provide a basis for understanding the effects of sperm ROS generation on zona pellucida penetration in male reproductive toxicology.

Acrosome Reaction↗
PubMed1999

Role of synaptic vesicle proton gradient and protein phosphorylation on ATP-mediated activation of membrane-associated brain glutamate decarboxylase.

Previously, we have shown that the soluble form of brain glutamic acid decarboxylase (GAD) is inhibited by ATP through protein phosphorylation and is activated by calcineurin-mediated protein dephosphorylation (Bao, J., Cheung, W. Y., and Wu, J. Y. (1995) J. Biol. Chem. 270, 6464-6467). Here we report that the membrane-associated form of GAD (MGAD) is greatly activated by ATP, whereas adenosine 5'-[beta,gamma-imido]triphosphate (AMP-PNP), a non-hydrolyzable ATP analog, has no effect on MGAD activity. ATP activation of MGAD is abolished by conditions that disrupt the proton gradient of synaptic vesicles, e.g. the presence of vesicular proton pump inhibitor, bafilomycin A1, the protonophore carbonyl cyanide m-chorophenylhydrazone or the ionophore gramicidin, indicating that the synaptic vesicle proton gradient is essential in ATP activation of MGAD. Furthermore, direct incorporation of (32)P from [gamma-(32)P]ATP into MGAD has been demonstrated. In addition, MGAD (presumably GAD65, since it is recognized by specific monoclonal antibody, GAD6, as well as specific anti-GAD65) has been reported to be associated with synaptic vesicles. Based on these results, a model linking gamma-aminobutyric acid (GABA) synthesis by MGAD to GABA packaging into synaptic vesicles by proton gradient-mediated GABA transport is presented. Activation of MGAD by phosphorylation appears to be mediated by a vesicular protein kinase that is controlled by the vesicular proton gradient.

Adenosine Triphosphate↗
PubMed1999

Echo-enhancing sonography of a large-vessel hemangioma of the neck.

Because of the slow flow in the venous spaces of large-vessel hemangiomas, demonstration of color flow signals with conventional color Doppler or power Doppler sonography may be difficult. We report the case of a 22-year-old female patient with a soft tissue tumor containing multiple fluid-filled spaces in the right supraclavicular region. Gray-scale, color, and power Doppler sonography could not differentiate between cystic lymphangioma and large-vessel hemangioma. The intravenous echo-enhancing contrast agent Levovist was administered, and a significant echo-enhancing effect on color and power Doppler imaging was demonstrated in the fluid-filled spaces and lasted for about 3 minutes. Histopathologic study of the excised tumor confirmed the sonographic diagnosis of hemangioma. Levovist appears useful in depicting slow flow in a large-vessel hemangioma.

Adult↗
PubMed1999

Increased cutaneous blood flow but impaired post-ischemic response of nutritional flow in obese children.

It is well known that cardiovascular risk factors and dyslipidemia along with obesity can be identified early in life. Serum lipids and lipoproteins influence skin vessel reactivity in adults. The purpose of this study was to investigate the cutaneous microcirculation and its correlation to dyslipidemia in obese children. Thirty-four obese children and an equal number of age- and sex-matched healthy children were enrolled in this study. Laser Doppler flowmetry and dynamic capillaroscopy were performed to measure total cutaneous blood flow and nutritional blood flow, respectively. A significantly higher cutaneous baseline flow and peak flow after a 1-min arterial occlusion was noticed in obese group as compared to normal control. Results of dynamic capillaroscopy revealed values of resting capillary blood cell velocity that did not differ from controls but a significantly lower peak CBV after a 1-min arterial occlusion in obesity. Plasma lipids examination showed a high triglycerides and low high density lipoprotein cholesterol levels in obesity. Pearson analysis detected no significant correlations between pCBV, baseline flow and peak flow and dyslipidemia in obesity. These results may be explained by the fact that the duration of dyslipidemia in our study population was not long enough to cause vascular damage or the dyslipidemia was not pronounced enough to have an impact on skin vessel activity tests. It is also possible, that the combination of methods used in the present study was not sensitive enough to detect an existing correlations between flow values and high triglycerides/low HDL-cholesterol values in the obese group. Our results suggest that all the responses of the cutaneous microcirculation were mainly due to physiological compensatory rather than pathological reactions in young obese children. The results of cutaneous microcirculatory assessment in obese children provide follow-up parameters for assessing the risk of cardiovascular diseases later in life.

Adolescent↗
PubMed1999

On the structural preservation of recombinant human growth hormone in a dried film of a synthetic biodegradable polymer.

In this work we describe the structural investigation of the model protein recombinant human growth hormone (rhGH) under conditions relevant to polymeric sustained-delivery depots, including the dried protein entrapped in a film of poly(DL-lactic-co-glycolic)acid. At each step of the procedure, dehydration of rhGH by lyophilization, suspension in methylene chloride, and drying from that suspension, the structure of rhGH was probed noninvasively using Fourier transform infrared (FTIR) spectroscopy. We found that the structure of rhGH was significantly changed by the dehydration process as indicated by a marked drop in the alpha-helix content and increase in the beta-sheet content. Subsequent suspension of this powder in methylene chloride, drying from that suspension, and drying from a methylene chloride/PLGA solution introduced only minor additional structural changes when using appropriate conditions. This result is likely due to the limited molecular mobility of proteins in nonprotein-dissolving organic solvents. Finally, when rhGH was co-lyophilized with the lyoprotectant trehalose, which preserves the secondary structure, the rhGH entrapped in the PLGA matrix also had a nativelike secondary structure.

Freeze Drying↗
PubMed1999

Influence of calcium ions on the structure and stability of recombinant human deoxyribonuclease I in the aqueous and lyophilized states.

The effect of calcium ions on the structure and stability of recombinant human DNase I (rhDNase) in the aqueous and solid (lyophilized) states was investigated. Fourier transform infrared (FTIR) spectroscopy was used to examine the overall secondary structure, while chemical stability was monitored in terms of deamidation and soluble aggregate formation at 40 degrees C. The exogenous calcium was removed by EGTA. This process can remove all but approximately one calcium ion per protein molecule. Analysis of the FTIR spectra in the amide III region in either the aqueous or lyophilized state demonstrated that removal of exogenous Ca2+ by EGTA-treatment had little effect on the secondary structure (and lyophilization-induced rearrangement thereof). For the aqueous solution, circular dichroism was used as an independent technique and confirmed that there was no large overall change in the secondary or tertiary structure upon the removal of calcium. The primary degradation route for the aqueous protein was deamidation. For the EGTA-treated protein, there was also severe covalent aggregation, e.g., formation of intermolecular disulfides facilitated by the cleavage of Cys173-Cys209. The aggregates exhibited a markedly different secondary structure compared to the native protein. For instance, the beta-sheet band observed at ca. 1620 cm-1 wavenumber in the amide I second derivative spectra was increased. Enzymatic activity was completely lost upon aggregation, consistent with the cleavage of the aforementioned native disulfide. For the protein lyophilized in the presence of Ca2+, there was no increase in deamidated species during solid-state storage; however, some aggregation was observed. For the lyophilized EGTA-treated protein, aggregation was even more pronounced, and there was some loss in enzymatic activity upon reconstitution. Thus, the removal of calcium ions by EGTA-treatment decreased the stability of rhDNase in both the aqueous and solid states even though no large overall calcium-induced structural changes could be observed by the techniques used in this study.

Calcium↗
PubMed1999

Lyophilization of protein formulations in vials: investigation of the relationship between resistance to vapor flow during primary drying and small-scale product collapse.

During the lyophilization process, formulations containing protein, bulking agent, or lyoprotectant form a dry product layer that can affect the transport of sublimed water vapor. We carried out an investigation of the primary drying segment of lyophilization to evaluate the relationship between the resistance to water vapor flow through the dried layer and the microstructure of the dried cake. Recombinant humanized antibody HER2 (rhuMAb HER2) formulated in trehalose was studied, as were protein-free formulations containing trehalose and sucrose. Sublimation rate and product temperature data were used to compute the resistance to mass transfer. Dried cake structure was examined by scanning electron microscopy and a novel fluorescence microscopy method. Collapse temperatures were determined by freeze-drying microscopy. Mass transfer resistance was found to decrease with increases in temperature for each material. Resistance also depended on composition, decreasing in the formulation series, rhuMAb HER2, trehalose, sucrose. The lyophilized material consisted of porous cakes, with a distinct denser region at the top. Formulation and temperature affected the microstructure of the dried cakes. The formulated trehalose and sucrose were seen, by both microscopy techniques, to possess small (2-20 microm) holes in their platelike structures after lyophilization. The quantity of holes was higher for material dried at higher temperature. The collapse temperature (Tc) of a material appeared to play a role in the process, as lower Tc was correlated with lower resistance and a greater extent of holes. Our results are consistent with the theory that lower resistance to water vapor flow in the primary drying stage of lyophilization may be due to small-scale product collapse.

Freeze Drying↗
PubMed1999

The effect of formulation excipients on protein stability and aerosol performance of spray-dried powders of a recombinant humanized anti-IgE monoclonal antibody.

PURPOSE: To study the effect of trehalose, lactose, and mannitol on the biochemical stability and aerosol performance of spray-dried powders of an anti-IgE humanized monoclonal antibody. METHODS: Protein aggregation of spray-dried powders stored at various temperature and relative humidity conditions was assayed by size exclusion chromatography and sodium dodecyl sulfate polyacrylamide gel electrophoresis. Protein glycation was determined by isoelectric focusing and affinity chromatography. Crystallization was examined by X-ray powder diffraction. Aerosol performance was assessed as the fine particle fraction (FPF) of the powders blended with coarse carrier lactose, and was determined using a multiple stage liquid impinger. RESULTS: Soluble protein aggregation consisting of non-covalent and disulfide-linked covalent dimers and trimers occurred during storage. Aggregate was minimized by formulation with trehalose at or above a molar ratio in the range of 300: 1 to 500:1 (excipient:protein). However, the powders were excessively cohesive and unsuitable for aerosol administration. Lactose had a similar stabilizing effect, and the powders exhibited acceptable aerosol performance, but protein glycation was observed during storage. The addition of mannitol also reduced aggregation, while maintaining the FPF, but only up to a molar ratio of 200:1. Further increased mannitol resulted in crystallization, which had a detrimental effect on protein stability and aerosol performance. CONCLUSIONS: Protein stability was improved by formulation with carbohydrate. However, a balance must be achieved between the addition of enough stabilizer to improve protein biochemical stability without compromising blended powder aerosol performance.

Aerosols↗