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Biomedical subjects

Y Zhao

Publications and source records attributed to Y Zhao.

At least 181 records · Page 10Linked to original sources
PubMed2001

AP-3 mediates tyrosinase but not TRP-1 trafficking in human melanocytes.

Patients with Hermansky-Pudlak syndrome type 2 (HPS-2) have mutations in the beta 3A subunit of adaptor complex-3 (AP-3) and functional deficiency of this complex. AP-3 serves as a coat protein in the formation of new vesicles, including, apparently, the platelet's dense body and the melanocyte's melanosome. We used HPS-2 melanocytes in culture to determine the role of AP-3 in the trafficking of the melanogenic proteins tyrosinase and tyrosinase-related protein-1 (TRP-1). TRP-1 displayed a typical melanosomal pattern in both normal and HPS-2 melanocytes. In contrast, tyrosinase exhibited a melanosomal (i.e., perinuclear and dendritic) pattern in normal cells but only a perinuclear pattern in the HPS-2 melanocytes. In addition, tyrosinase exhibited a normal pattern of expression in HPS-2 melanocytes transfected with a cDNA encoding the beta 3A subunit of the AP-3 complex. This suggests a role for AP-3 in the normal trafficking of tyrosinase to premelanosomes, consistent with the presence of a dileucine recognition signal in the C-terminal portion of the tyrosinase molecule. In the AP-3-deficient cells, tyrosinase was also present in structures resembling late endosomes or multivesicular bodies; these vesicles contained exvaginations devoid of tyrosinase. This suggests that, under normal circumstances, AP-3 may act on multivesicular bodies to form tyrosinase-containing vesicles destined to fuse with premelanosomes. Finally, our studies demonstrate that tyrosinase and TRP-1 use different mechanisms to reach their premelanosomal destination.

Adaptor Proteins, Vesicular Transport↗
PubMed2001

Differential expression of TGF-beta1 and TGF-beta3 in serosal tissues of human intraperitoneal organs and peritoneal adhesions.

Elevated local expression of transforming growth factor (TGF-beta) has been associated with increased incidence of peritoneal adhesion formation. In this study we determine whether differences in basal expression of TGF-beta in serosal tissue of peritoneal organs correlate with incidence of adhesion formation. Serosal tissue of parietal peritoneum, uterus, oviduct, ovary, omentum, large and small bowels as well as adhesions, skin, fascia, subcutaneous tissue, peritoneal fluid and serum were collected from 57 subjects with/without adhesions who were undergoing abdominal/pelvic surgery. To determine TGF-beta1 and TGF-beta3 mRNA and protein expression, total RNA and protein were isolated from these tissues and along with the fluids, subjected to quantitative RT-PCR and enzyme-linked immunosorbent assay (ELISA) respectively. Tissue sections were immunostained for TGF-beta1 and TGF-beta3 protein. We found that TGF-beta1 and TGF-beta3 mRNA and protein are expressed in these tissues and present in peritoneal fluids and serum, with considerable variations in level of their expression. Comparatively, there was more variation in TGF-beta1 than TGF-beta3 expression without age or gender relation. Adhesions express a significantly higher TGF-beta1 mRNA and have the highest TGF-beta1:TGF-beta3 ratio, with lowest concentrations and ratio detected in omentum, small and large bowels; in contrast uterus expresses higher TGF-beta3, with lowest concentrations detected in subcutaneous tissue and large bowels (P < 0.05). A similar trend was also observed for total (active + latent) TGF-beta1 protein expression, with low active TGF-beta1 that was not significantly different among the tissue extracts and fluids. However, the lowest active:total TGF-beta1 ratio was found in adhesions and ovary. In subjects with adhesions, the adhesions express significantly more TGF-beta1 compared to parietal peritoneum (P < 0.05). Immunoreactive TGF-beta1 and TGF-beta3 protein were present in various cell types in these tissues with intensity reflecting their mRNA and protein expression. In conclusion, we provided evidence that serosal tissue of various peritoneal organs and adhesions express TGF-beta1 and TGF-beta3. Since TGF-beta is expressed differently in these tissues and tissue injury often alters the expression of TGF-beta, we propose that tissues with a higher basal expression of TGF-beta may become predisposed to develop more adhesions compared to others.

Adult↗
PubMed2001

Leukocyte rolling in rat mesentery venules: distribution of adhesion bonds and the effects of cytoactive agents.

A new method of analyzing in vivo measurements of leukocyte WBC rolling along venular endothelium (EC) has been developed to extract insightful information on the dynamics of WBC-EC bond formation and disruption. The rolling velocity of WBCs was obtained by intravital microscopy of rat mesenteric venules. For the "spontaneous" rolling observed following exteriorization of the mesentery, we estimated that the average distance between clusters of adhesion bonds which tether a rolling cell to the venular wall was about 2 microm, and that the average lifetime of a bond cluster at the trailing edge of the rolling cell, from its exposure to the tensile force to its release, was on the order of 0.05 s. Both the inter-cluster distance and the lifetime were significantly reduced by treatments with the chemoattractant N-formyl-methionyl-leucyl-phenylalanine and the cytokine interleukin-1, while the average lifetime of the stretched bond clusters was not significantly changed by treatment with the cytoskeleton-modifying agents cytochalasin B and colchicine. Each of the four treatments significantly reduced the heterogeneity in the cell rolling velocity, presumably by the selective recruitment of WBC subsets from the circulating WBC population or by a reduction in the heterogeneity of endothelial adhesiveness. These results were analyzed in the context of in vitro data in the literature on molecular bonds of cell adhesion. The findings suggest that, in the case of "spontaneous" rolling, there are on average approximately 2-3 clusters of adhesion bonds between a rolling cell and the vessel wall, and approximately five bonds in each cluster.

Animals↗
PubMed2001

Altered extracellular signal-regulated kinase signaling and glycogen metabolism in skeletal muscle from p90 ribosomal S6 kinase 2 knockout mice.

The p90 ribosomal S6 kinase (RSK), a cytosolic substrate for the extracellular signal-regulated kinase (ERK), is involved in transcriptional regulation, and one isoform (RSK2) has been implicated in the activation of glycogen synthase by insulin. To determine RSK2 function in vivo, mice lacking a functional rsk2 gene were generated and studied in response to insulin and exercise, two potent stimulators of the ERK cascade in skeletal muscle. RSK2 knockout (KO) mice weigh 10% less and are 14% shorter than wild-type (WT) mice. They also have impaired learning and coordination. Hindlimb skeletal muscles were obtained from mice 10, 15, or 30 min after insulin injection or immediately after strenuous treadmill exercise for 60 min. While insulin and exercise significantly increased ERK phosphorylation in skeletal muscle from both WT and KO mice, the increases were twofold greater in the KO animals. This occurred despite 27% lower ERK2 protein expression in skeletal muscle of KO mice. KO mice had 18% less muscle glycogen in the fasted basal state, and insulin increased glycogen synthase activity more in KO than WT mice. The enhanced insulin-stimulated increases in ERK and glycogen synthase activities in KO mice were not associated with higher insulin receptor or with IRS1 tyrosine phosphorylation or with IRS1 binding to phosphatidylinositol 3-kinase. However, insulin-stimulated serine phosphorylation of Akt was significantly higher in the KO animals. c-fos mRNA was increased similarly in muscle from WT and KO mice in response to insulin (2. 5-fold) and exercise (15-fold). In conclusion, RSK2 likely plays a major role in feedback inhibition of the ERK pathway in skeletal muscle. Furthermore, RSK2 is not required for activation of muscle glycogen synthase by insulin but may indirectly modulate muscle glycogen synthase activity and/or glycogen content by other mechanisms, possibly through regulation of Akt. RSK2 knockout mice may be a good animal model for the study of Coffin-Lowry syndrome.

Animals↗
PubMed2001

Distinct RNP complexes of shuttling hnRNP proteins with pre-mRNA and mRNA: candidate intermediates in formation and export of mRNA.

Nascent pre-mRNAs associate with hnRNP proteins in hnRNP complexes, the natural substrates for mRNA processing. Several lines of evidence indicate that hnRNP complexes undergo substantial remodeling during mRNA formation and export. Here we report the isolation of three distinct types of pre-mRNP and mRNP complexes from HeLa cells associated with hnRNP A1, a shuttling hnRNP protein. Based on their RNA and protein compositions, these complexes are likely to represent distinct stages in the nucleocytoplasmic shuttling pathway of hnRNP A1 with its bound RNAs. In the cytoplasm, A1 is associated with its nuclear import receptor (transportin), the cytoplasmic poly(A)-binding protein, and mRNA. In the nucleus, A1 is found in two distinct types of complexes that are differently associated with nuclear structures. One class contains pre-mRNA and mRNA and is identical to previously described hnRNP complexes. The other class behaves as freely diffusible nuclear mRNPs (nmRNPs) at late nuclear stages of maturation and possibly associated with nuclear mRNA export. These nmRNPs differ from hnRNPs in that while they contain shuttling hnRNP proteins, the mRNA export factor REF, and mRNA, they do not contain nonshuttling hnRNP proteins or pre-mRNA. Importantly, nmRNPs also contain proteins not found in hnRNP complexes. These include the alternatively spliced isoforms D01 and D02 of the hnRNP D proteins, the E0 isoform of the hnRNP E proteins, and LRP130, a previously reported protein with unknown function that appears to have a novel type of RNA-binding domain. The characteristics of these complexes indicate that they result from RNP remodeling associated with mRNA maturation and delineate specific changes in RNP protein composition during formation and transport of mRNA in vivo.

Animals↗
PubMed2001

Erythropoietin acts as a trophic factor in neonatal rat intestine.

BACKGROUND: Erythropoietin (Epo) receptors are present on enterocytes of fetal and neonatal small bowel but the role of Epo in the bowel is not known. AIMS: We tested the following hypotheses: (1) enterally dosed Epo is absorbed from the intestines of neonatal rats, (2) Epo acts as a trophic factor in developing small bowel, and (3) the trophic effects of Epo are dependent on the route of administration. METHODS: The dose dependent effects of enterally dosed recombinant human erythropoietin (rEpo 0--1000 U/kg/day) were studied in artificially raised rat pups and compared with dam raised controls and dam raised pups given rEpo in rat milk. After one week, reticulocyte counts, haematocrits, and plasma Epo concentrations were measured, and calibrated morphometric measurements of villi were performed. The effects of route of rEpo administration (enteral v parenteral) on erythropoiesis, bowel growth, and disaccharidase activity were studied in nursing pups treated for one and two weeks. RESULTS: Serum Epo concentrations ranged from undetectable (<0.6 mU/ml) to 8.4 mU/ml in control and enterally dosed pups (median 1.8 mU/ml), and from 4.9 to 82.3 mU/ml (median 20.4 mU/ml) in parenterally dosed animals. No increase in haematocrit or reticulocyte count was noted in enterally treated pups compared with controls after up to two weeks of treatment. Small bowel length was greater in rEpo treated pups, and a dose dependent increase in villus surface area which was independent of the route of dosing and associated with increased BrdU uptake was found. CONCLUSIONS: rEpo is not enterally absorbed in an intact and functional form from the intestines of neonatal rat pups. Thus enterally dosed rEpo has no erythropoietic effects. However, rEpo acts as a trophic factor in developing rat small bowel whether given enterally or parenterally.

Animals↗
PubMed2001

Assembly and activation of HK-PK complex on endothelial cells results in bradykinin liberation and NO formation.

Prekallikrein (PK) activation on human umbilical endothelial cells (HUVEC) presumably leads to bradykinin liberation. On HUVEC, PK activation requires the presence of cell-bound high-molecular-weight kininogen (HK) and Zn(2+). We examined the Zn(2+) requirement for HK binding to and the consequences of PK activation on endothelial cells. Optimal HK binding (14 pmol/10(6) HUVEC) is seen with no added Zn(2+) in HEPES-Tyrode buffer containing gelatin versus 16--32 microM added Zn(2+) in the same buffer containing bovine serum albumin. The affinity and number of HK binding sites on HUVEC are a dissociation constant of 9.6 +/- 1.8 nM and a maximal binding of 1.08 +/- 0.26 x 10(7) sites/cell (means +/- SD). PK is activated to kallikrein by an antipain-sensitive mechanism in the presence of HK and Zn(2+) on HUVEC, human microvascular endothelial cells, umbilical artery smooth muscle cells, and bovine pulmonary artery endothelial cells. Simultaneous with kallikrein formation, bradykinin (5.0 or 10.3 pmol/10(6) HUVEC in the absence or presence of lisinopril, respectively) is liberated from cell-bound HK. Liberated bradykinin stimulates the endothelial cell bradykinin B2 receptor to form nitric oxide. Assembly and activation of PK on endothelial cells modulates their physiological activities.

Animals↗
PubMed2001

Lack of mitochondrial trifunctional protein in mice causes neonatal hypoglycemia and sudden death.

Mitochondrial trifunctional protein (MTP) is a hetero-octamer of four alpha and four beta subunits that catalyzes the final three steps of mitochondrial long chain fatty acid beta-oxidation. Human MTP deficiency causes Reye-like syndrome, cardiomyopathy, or sudden unexpected death. We used gene targeting to generate an MTP alpha subunit null allele and to produce mice that lack MTP alpha and beta subunits. The Mtpa(-/-) fetuses accumulate long chain fatty acid metabolites and have low birth weight compared with the Mtpa(+/-) and Mtpa(+/+) littermates. Mtpa(-/-) mice suffer neonatal hypoglycemia and sudden death 6-36 hours after birth. Analysis of the histopathological changes in the Mtpa(-/-) pups revealed rapid development of hepatic steatosis after birth and, later, significant necrosis and acute degeneration of the cardiac and diaphragmatic myocytes. This mouse model documents that intact mitochondrial long chain fatty acid oxidation is essential for fetal development and for survival after birth. Deficiency of MTP causes fetal growth retardation, neonatal hypoglycemia, and sudden death.

Animals↗
PubMed2001

The novel gene locus for agenesis of permanent teeth (He-Zhao deficiency) maps to chromosome 10q11.2.

He-Zhao deficiency has been recently characterized with a distinct form of agenesis of permanent teeth that is different from other previously reported disorders of tooth agenesis. This inherited abnormality suggests that some gene(s) associated with the development of permanent teeth may mutate. In this study, we map the gene locus to chromosome 10q11.2. The DNA pooling method combined with two-point and multi-point linkage analysis has been successfully applied. The maximum LOD (Zmax) scores for two-point and multi-point analyses are 13.29 (on marker D10S196) at recombination fraction (theta) = 0 and 18.09 (between markers D10S1772 and D10S1766), respectively. Haplotype analysis confined the locus within an interval of 5.5 cM flanked by markers D10S604 and D10S568. This study has demonstrated a novel gene locus responsible for He-Zhao deficiency and provides a good likelihood for the discovery of one of the genes determining permanent tooth formation and development.

Alleles↗
PubMed2001

Two Chinese families with Pendred's syndrome--radiological imaging of the ear and molecular analysis of the pendrin gene.

We report two families in whom the index cases satisfied the classical diagnostic criteria of Pendred's syndrome. In family I, two siblings were deaf, and one was normal. In family II, both parents and two offspring were deaf. Computed tomography scans performed in five of six of these deaf individuals showed enlarged vestibular aqueducts in all cases, and Mondini cochlea only in family II. Affected members in family I were compound heterozygotes inheriting the paternal allele with a novel mutation S398del in exon 10 and a maternal allele with two mutations IVS13+9C-->G in intron 13, in addition to H723R. In family II, the mother and one child carried both the novel intronic IVS8-2A-->G and H723R mutations, whereas the father and index case were homozygous for the IVS8-2A-->G mutation. A perchlorate discharge test was positive in 50% of cases tested. In conclusion, we concur that radiological and molecular studies should be performed for confirmation of Pendred's syndrome. We report, for the first time, a Pendred's syndrome family in which affected members had three mutations, as well as a second family in whom the intermarriage of two Pendred's syndrome patients resulted in Pendred's syndrome offspring.

Adolescent↗
PubMed2001

Functional specificity of the Hoxa13 homeobox.

To better define Abd-B type homeodomain function, to test models that predict functional equivalence of all Hox genes and to initiate a search for the downstream targets of Hoxa13, we have performed a homeobox swap by replacing the homeobox of the Hoxa11 gene with that of the Hoxa13 gene. The Hoxa11 and Hoxa13 genes are contiguous Abd-B type genes located at the 5' end of the HoxA cluster. The modified Hoxa11 allele (A11(13hd)) showed near wild-type function in the development of the kidneys, axial skeleton and male reproductive tract, consistent with functional equivalence models. In the limbs and female reproductive tract, however, the A11(13hd) allele appeared to assume dominant Hoxa13 function. The uterus, in particular, showed a striking homeotic transformation towards cervix/vagina, where Hoxa13 is normally expressed. Gene chips were used to create a molecular portrait of this tissue conversion and revealed over 100 diagnostic gene expression changes. This work identifies candidate downstream targets of the Hoxa13 gene and demonstrates that even contiguous Abd-B homeoboxes have functional specificity.

Alleles↗
PubMed2001

Inhibitory effects of RRR-alpha-tocopheryl succinate on benzo(a)pyrene (B(a)P)-induced forestomach carcinogenesis in female mice.

AIM: To study the inhibitory effects of VES (RRR-alpha-tocopheryl Succinate, VES),a derivative of natural Vitamin E, on benzo(a)pyrene(B(a)P)-induced forestomach tumor in female mice. METHODS: The model of B(a)P-induced forestomach tumor was established according to the methods of Wattenberg with slight modify-cations. One hundred and eighty female mice (6 weeks old) were divided into six groups equally; negative control (Succinic acid), vehicle control (Succinate+B(a)P),positive control(B(a)P), high VES(2.5 g/kg.b.w+B(a)P), low VES(1.25 g/kg.b.w+B(a)P)ig as well as VES by ip (20 mg/kg.b.w+B(a)P). Except the negative control group, the mice were administrated with B(a)P ig. and corresponding treatments for 4 weeks to study the anti-carcinogenetic effect of VES during the initiation period. The experiment lasted 29 weeks, in which the inhibitory effects of VES both on tumor incidence and tumor size were tested. RESULTS: The models of B(a)P-induced forestomach tumor in female mice were established successfully. Some were cauliflower-like, others looked like papilla, even a few were formed into the ulcer cavities. VES at 1.25 g/kg.b.w, 2.5 g/kg.b.w. by ig and 20 mg/kg.b.w. via ip could decrease the number of tumors per mouse (1.7 plus minus 0.41, 1.6 plus minus 0.34 and 1.1 +/- 0.43), being lower than that of B(a)P group (5.4 +/- 0.32, P<0.05). The tumor incidence was inhibited by 18.2%, 23.1% and 50.0%. VES at 1.25 g/kg.b.w., 2.5 g/kg.b.w. by ig and 20 mg/kg.b.w. via ip reduced the total volume of tumors per mouse (54.8 +/- 8.84, 28.4 8 +/- 8.32 and 23.9 8 +/- 16.05), being significantly lower than that of B(a)P group (150.2 8 +/- 20.93, P < 0.01). The inhibitory rates were 63.5%, 81.1% and 84.1%, respectively. CONCLUSION: VES has inhibitory effects on B(a)P-induced forestomach carcinogenesis in female mice, especially by ip and it may be a potential anti-cancer agent in vivo.

Animals↗
PubMed2001

Distribution of nitric oxide synthase in stomach myenteric plexus of rats.

AIM: To study the distribution of nitric oxide synthase (NOS) in rat stomach myenteric plexus. METHODS: The distribution of NOS in gastric wall was studied in quantity and location by the NADPH-diaphorase (NDP) histochemical staining method and whole mount preparation technique. RESULTS: NOS was distributed in whole stomach wall, most of them were located in myenteric plexus, and distributed in submucosal plexus.The shape of NOS positive neurons was basically similar, most of them being round and oval in shape. But their density, size and staining intensity varied greatly in the different parts of stomach. The density was 62+/-38 cells mm(2) (antrum), 43+/-32 cells/mm(2) (body), and 32+/-28 cells mm(2) (fundus), respectively. The size and staining intensity of NOS positive neurons in the fundus were basically the same, the neurons being large and dark stained, while they were obviously different in antrum. In the body of the stomach, the NOS positive neurons were in an intermediate state from fundus to antrum. There were some beadlike structures which were strung together by NOS positive varicosities in nerve fibers, some were closely adherent to the outer walls of blood vessels. CONCLUSION: Nitric oxide might be involved in the modulation of motility, secretion and blood circulation of the stomach, and the significant difference of NOS positive neurons in different parts of stomach myenteric plexus may be related to the physiologic function of stomach.

Animals↗
PubMed2001

Ultrasonographic texture analysis of parenchymatous organs by the four-neighborhood-pixels algorithm: clinical experiment.

OBJECTIVE: The parenchyma of organs such as liver, thyroid, and mammary gland during climacterium have common ultrasonographic textural features, which together form what we call small-dot-structure texture. To study this texture we designed the 4-neighborhood-pixels algorithm, an ultrasonographic texture analysis algorithm. The objective of this study was to confirm whether the 4-neighborhood-pixels algorithm can reflect the features of small-dot-structure texture. METHODS: A changed small-dot-structure texture and 3 other textures were compared with the normal small-dot-structure texture in 4 groups, and a histogram algorithm was used for contrast with the 4-neighborhood-pixels algorithm. RESULTS: The 4-neighborhood-pixels algorithm could reflect all the textural differences, but the histogram algorithm could reflect only some of them. CONCLUSIONS: The 4-neighborhood-pixels algorithm is a good algorithm for analyzing ultrasonographic small-dot-structure texture. Not only can it reflect changes in the small-dot-structure texture, but it can also differentiate between small-dot-structure and non-small-dot-structure textures.

Adipose Tissue↗
PubMed2001

[Analysis of the non-deletion alpha-thalassemia mutations by PCR temperature gradient gel electrophoresis].

OBJECTIVE: To develop a PCR-temperature gradient gel electrophoresis(TGGE) method applied to screening for the point mutations causing the non-deletion alpha-thalassemia. METHODS: The entire alpha2-globin gene fragment (1085 bp) was selectively amplified from human genomic DNA with different genotypes of alpha-thalassemia and a 543 bp fragment spanning the exon 2 and exon 3 of the alpha2-globin gene was amplified with a pair of nested primers. The condition of perpendicular and parallel TGGE for the two different fragments in length was optimized and the candidate mutant was confirmed by DNA sequencing. In pilot study, 15 samples with suspected non-deletion alpha-thalassemia were screened for point mutations in alpha-globin gene. RESULTS: Hb Constant Spring (Hb CS) and Hb Quong Sze (Hb QS), two most commonly types of the non-deletion alpha-thalassemia distributed in Chinese, and a rare type of the Hb Westmead mutation, alpha2 CD122 CAC-->CAG(His-->Gln) could be detected by PCR-TGGE. Among those 15 samples screened for alpha-thalassemia mutation were 10 samples with Hb CS mutation, 2 samples with Hb QS mutation, 2 samples with Hb Westmead mutation, and a previously unreported one, alpha2 CD31 AGG-->AAG(Arg-->Lys), confirmed by DNA sequencing. CONCLUSION: The present PCR-TGGE method could be a useful tool for the molecular screening for the point mutations causing alpha-thalassemia and alpha-globin gene polymorphism.

Electrophoresis↗