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X Ding

Publications and source records attributed to X Ding.

At least 127 records · Page 7Linked to original sources
PubMed1997

cDNA cloning, heterologous expression, and characterization of mouse CYP2G1, an olfactory-specific steroid hydroxylase.

CYP2G1 is expressed specifically in the olfactory mucosa in rabbits and rats. In the present study, a full-length cDNA for mouse CYP2G1 was obtained using a PCR approach with RNA preparations from the olfactory mucosa of C57BL/6 mice. Sequence comparisons indicated that mouse CYP2G1 is highly homologous in deduced amino acid sequence to rabbit (82.4% identity) and rat CYP2G1 (94.9% identity). RNA blot and immunoblot analyses indicated that mouse CYP2G1 is expressed only in the olfactory mucosa. The coding region of the mouse CYP2G1 cDNA was cloned into a baculoviral expression vector for heterologous production of the enzyme in cultured insect cells. Heterologously expressed mouse CYP2G1 was active in a reconstituted system toward testosterone and progesterone, producing all the major metabolites detected in olfactory microsomal reactions, including 15 alpha-, 15 beta-, and 2 beta-hydroxytestosterone from testosterone and two unidentified metabolites from progesterone. Kinetic analysis indicated that mouse CYP2G1 has relatively high affinities toward the steroid substrates, with K(m) values in the micromolar range for both testosterone and progesterone. At a substrate concentration of 10 microM, microsomes of olfactory mucosa had much higher turnover numbers toward testosterone and progesterone than hepatic microsomes, consistent with the olfactory-specific expression of a high-affinity sex steroid hydroxylase. These findings will facilitate further molecular genetics studies on the biological function of CYP2G1 in a mouse model.

Amino Acid Sequence↗
PubMed1997

CDNA cloning, heterologous expression, and characterization of rat intestinal CYP2J4.

The small intestine is the major portal of entry of ingested xenobiotics. Previous studies from this and other laboratories indicated that at least 6 of the 33 xenobiotic metabolizing forms of P450 currently identified are expressed in rat small intestinal epithelial cells. In the present study, a previously unidentified rat P450, designated CYP2J4, was identified in rat small intestine using PCR. The full-length CYP2J4 cDNA contains an open reading frame for a protein of 501 residues and is 72.5 and 75.8% identical to rabbit CYP2J1 and human CYP2J2, respectively, in deduced amino acid sequences. The coding region of CYP2J4 cDNA has been cloned into a baculoviral expression vector (pVL1392) and expressed in cultured Spodoptera frugiperta (SF9) cells. The heterologously expressed CYP2J4 protein displayed a typical p450 CO-difference spectrum, with maximum absorbance at 449 nm. When purified to near electrophoretic homogeneity, it was active toward arachidonic acid in a reconstituted system with NADPH-P450 reductase and phospholipid, producing both hydroxyeicosatetraenoic and epoxyeicosatrienoic acids. RNA blot analysis with CYP2J4 cDNA as a probe detected two mRNA species, about 2.0 and 2.4 kb, respectively, in RNA preparations from liver, intestine, olfactory mucosa, kidney, heart, and lung. The 2.0-kb mRNA species was abundant in liver, small intestine, and olfactory mucosa, whereas the 2.4-kb mRNA species was predominant only in the olfactory mucosa. Immunoblot analysis of microsomal fractions from different rat tissues with a polyclonal anti-peptide antibody to CYP2J4 detected a protein with the same electrophoretic mobility as purified CYP2J4 most abundantly in small intestine and to a lesser extent in liver and other immunoreactive proteins with slightly higher electrophoretic mobility than purified CYP2J4 in a number of tissues, including small intestine, liver, kidney, lung, and olfactory mucosa. The predominant distribution of CYP2J4, which has activity toward arachidonic acid, is provocative, but its physiological function is as yet unknown.

Amino Acid Sequence↗
PubMed1997

Development and characterization of desmoglein-3 specific T cells from patients with pemphigus vulgaris.

Pemphigus vulgaris (PV) is a cutaneous autoimmune disease characterized by blister formation in the suprabasilar layers of skin and mucosae and anti-desmoglein-3 (Dsg3) autoantibodies bound to the surface of lesional keratinocytes and circulating in the serum of patients. This disease can be reproduced in neonatal mice by passive transfer of patients' IgG, indicating that humoral immunity plays an important role in the pathogenesis of PV. Currently, the role of T lymphocytes in the development of PV is not clear. Here, we report that three immunoreactive segments of the ectodomain of Dsg3 specifically induced proliferation of T cells from PV patients. We found that T lymphocytes from 13 out of 14 patients responded to at least one of three Dsg3 peptides. T cells from controls and other patient groups did not respond to these Dsg3 peptides. The major T cell population stimulated by these Dsg3 peptides was CD4 positive. Dsg3-specific T cell lines and clones were developed and were shown to express a CD4 positive memory T cell phenotype. Upon stimulation, these cell lines and clones secreted a Th2-like cytokine profile. The Dsg3 responses of these T cells were restricted to HLA-DR, and not -DQ and -DP, of the major histocompatibility complex. This information will help to elucidate the cellular immune abnormalities leading to production of pathogenic IgG autoantibodies in patients with PV.

Autoantibodies↗
PubMed1997

Mechanisms of acantholysis in pemphigus vulgaris: role of IgG valence.

Pemphigus vulgaris (PV) is a dermatosis mediated by autoantibodies against desmoglein 3 (Dsg3). It was known that intraperitoneal (i.p.) injections of PV IgG and PV F(ab')2, but not of PV Fab, into neonatal mice reproduced the key features of the disease in these animals. It was proposed that crosslinking of antigen by bivalent PV autoantibodies may trigger acantholysis in PV. In the present study, we have used subcutaneous (s.c.) injections of PV IgG and its proteolytic fragments into neonatal mice to test equimolar amounts of these autoantibody fractions. Mice developed clinical and histological features of PV in a dose-dependent manner following a similar time course. PV IgG and Fab fractions induced acantholysis as early as 2 hr after the injection. It was also demonstrated that sc injections of PV Fab were more effective in inducing disease than ip injections. PV autoantibodies may bind an "adhesive site" of Dsg3 and impair its function, thus triggering acantholysis.

Acantholysis↗
PubMed1997

Nasal cytochrome P450 2A: identification, regional localization, and metabolic activity toward hexamethylphosphoramide, a known nasal carcinogen.

Two members of the cytochrome P450 2A subfamily, CYP2A10 and 2A11, are abundant nasal enzymes previously characterized in rabbit olfactory microsomes. Rabbit CYP2A is active toward a number of nasal toxicants, including the rat nasal procarcinogen hexamethylphosphoramide (HMPA). While P450s immunochemically related to the rabbit CYP2As have been detected in rat and human nasal mucosa, confirmation of these enzymes as members of the CYP2A subfamily and efforts to characterize their ability to bioactivate toxicants have been limited. In the present study, the regional distribution and cell-specific expression of CYP2A in the rat nasal cavity were examined using an antibody to rabbit CYP2A10/11. In sections of the anterior nose, immunoreactive CYP2A was present in ciliated cells of the nasal respiratory epithelium and cuboidal epithelial cells of the nasal transitional epithelium, but was absent in squamous epithelial cells. The most intense immunostaining was observed in the posterior nose. Olfactory sustentacular cells and Bowman's gland cells in sections posterior to the nasal papilla stained most intensely. Western blot analysis revealed that anti-CYP2A10/11 recognized a sharp band of approximately 50 kDa in nasal respiratory and olfactory microsomes, supporting the premise that the antibody is reacting with a cytochrome P450 enzyme. The nasal expression of CYP2A6 mRNA--a member of the human CYP2A subfamily having a high degree of homology to rabbit 2A10 and 2A11--was examined in human surgical patients. Middle turbinectomy tissues--largely composed of nasal respiratory epithelia--from 11 patients were analyzed for the presence of CYP2A6 using reverse transcription-polymerase chain reaction (RT-PCR). Identification of CYP2A6 was confirmed by DNA sequencing of RT-PCR products. CYP2A6 mRNA was detected in all of the human samples analyzed. In additional experiments, human CYP2A6 metabolized HMPA to formaldehyde, suggesting that this compound might cause nasal toxicity in humans. The identification of CYP2A cytochromes in rat and human nasal tissues may have important implications for risk assessment of inhaled xenobiotics.

Animals↗
PubMed1997

Intraperitoneal administration of coumarin causes tissue-selective depletion of cytochromes P450 and cytotoxicity in the olfactory mucosa.

Coumarin is a naturally occurring fragrant compound widely used in consumer products and also as a therapeutic agent. The effects of intraperitoneal (ip) and oral administration of coumarin on cytochrome P450 (P450) expression in olfactory mucosa were examined. A single ip injection of coumarin at 50 mg/kg resulted in a significant reduction of levels of CYP2A and CYP2G in the olfactory mucosa of Wistar rats and C57BL/6 mice at 48 hr following injection. Dose-response analysis of coumarin effects indicated that Wistar rats were more sensitive than C57BL/6 mice. A significant suppression of nasal CYP2A levels was observed at 25 mg/kg in rats, but not in mice. Depletion of P450 content was not observed in liver of either rats or mice at 50 mg/kg, indicating tissue-selective effects. Decreased P450 levels were observed at 24 hr, 48 hr, and 7 days following treatment, with minimal levels seen at 48 hr. The decrease in P450 levels was accompanied by necrosis, cell loss, and basal cell metaplasia in the olfactory mucosa. Intraperitoneal injection of 7-hydroxycoumarin or 3,4-dihydrocoumarin at 50 mg/kg did not result in depletion of nasal P450, indicating that the toxicity is not mediated by P450-catalyzed coumarin 7-hydroxylation and supporting the hypothesis that the formation of coumarin 3,4-epoxide may be responsible for the toxicity. Oral treatment with coumarin in drinking water led to a small, yet significant induction of CYP2A protein and coumarin hydroxylase activity in the nasal mucosa of mice, but not rats. Thus, ip administration of coumarin causes tissue-selective depletion of P450 and cytotoxicity in the olfactory mucosa of Wistar rats and C57BL/6 mice. It remains to be determined whether similar toxicity occurs following coumarin administration by other routes.

Animals↗
PubMed1997

Ticks and tick-borne bovine piroplasmosis in the Southmountain pasture of Hunan Province, China.

Ticks and tick-borne bovine piroplasmosis is one of the most serious diseases affecting cattle bred in the Southmountain pasture of Hunan province. An investigation carried out from 1992 to 1995 showed that Theileria sergenti was found in the blood of disease-stricken cows. Ticks were collected from cattle. The species concerned (three genera and four species of ticks: Haemaphysalis vietnamensis, Haemaphysalis longicornis, Ixodes sinensis and Boophilus microplus) were identified by microscopy and their numbers recorded every month. Because H. longicornis is a three-host tick, with the larvae, nymphs and adults all infesting cows and seasonal fluctuations in nymphs and adults correlating with theileriosis outbreaks, this tick species was a potential vector of T. sergenti. Epidemiological studies indicated that this disease usually occurred in summer or autumn. The disease attacked calves from May to August reaching a peak in June or July (as happened from 1992-1994). In cattle, the disease occurred from June to September with a peak in September. The following pathological lesions were observed: haemorrhages were seen in the subcutaneous tissues and serosa, in the intestinal mucosa and the mesenteric lymph nodes. The surface of the abomasum looked like a piece of red cloth. The liver was enlarged, its edge blunt and curled over. The gall bladder was filled with tawny coloured bile. The spleen was enlarged and dark brown. The heart was enlarged with numerous haemorrhagic foci in the auricle. The blood clotted incompletely. Sick cattle were treated with Berenil and imidocarb and the death rate was reduced from 26% to 5.9%.

Animals↗
PubMed1997

Mucosal and mucocutaneous (generalized) pemphigus vulgaris show distinct autoantibody profiles.

Pemphigus vulgaris is an autoimmune bullous disorder characterized by autoantibodies directed against desmoglein 3. A group of 19 pemphigus vulgaris sera were characterized by immunoblotting, immunofluorescence, immunoprecipitation, and the passive transfer mouse model. The aim of these studies was to determine the specificity of the autoantibody response in these patients. All patients had clinical and histologic evidence of pemphigus vulgaris. Fogo selvagem sera (n = 8), bullous pemphigoid sera (n = 8), antinuclear antibodies positive sera from patients with lupus erythematosus (n = 2), and normal human sera (n = 8) were used as controls. All pemphigus vulgaris patients showed titers of IgG autoantibodies by indirect immunofluorescence > or = 1:60, predominantly of the IgG4 subclass and immunoprecipitated recombinant desmoglein 3 expressed in the baculovirus system. Patients with disease localized to the mucous membranes showed no reactivity with desmoglein 1 and only one had weak reactivity with mouse skin by indirect immunofluorescence (titer = 1:20). Sera of four of these mucosal patients were tested in the mouse model and three of four did not elicit skin or mucosal disease in the animals. In contrast, sera from all seven patients with disease involving the skin and mucous membranes (generalized disease) produced disease in neonatal mice. In one patient the disease evolved from pure mucosal involvement associated with anti-desmoglein 3 antibodies to a disorder involving mucosas and skin. This transition was associated with the appearance of anti-desmoglein 1 antibodies in the patient's serum. These studies indicate that the autoantibody response in pemphigus vulgaris is heterogeneous. Epitopes recognized by some pemphigus vulgaris sera are species specific and others may be mucosal specific.

Animals↗
PubMed1997

T lymphocytes from a subset of patients with pemphigus vulgaris respond to both desmoglein-3 and desmoglein-1.

Pemphigus vulgaris and pemphigus foliaceus are cutaneous autoimmune diseases characterized by intraepithelial blisters and autoantibodies to desmosomal glycoproteins. The antigens recognized by pemphigus vulgaris and pemphigus foliaceus autoantibodies are desmoglein-3 (Dsg3) and desmoglein-1 (Dsg1), respectively. Dsg3 and Dsg1 are members of the desmoglein subfamily of the cadherin supergene family of cell adhesion molecules. It has been well documented that a subset of pemphigus vulgaris sera have IgG reactivity to both Dsg1 and Dsg3, suggesting that Dsg1 may also participate in the autoimmune response of these patients. The cellular mechanisms of T cell autoimmunity in these patients, however, are completely unknown. In this study, we tested the proliferative responses of T lymphocytes from eight pemphigus vulgaris patients after incubation with Dsg3 and Dsg1 fusion proteins. The sera of four of these PV patients showed reactivity with both Dsg1 and Dsg3, whereas the remaining four reacted only with Dsg3. We found that T cells obtained from those patients that exhibited the combined Dsg1/Dsg3 autoantibody reactivity showed a proliferative response after exposure to either Dsg1 or Dsg3 fusion proteins. The cellular responses to both of these recombinant proteins were highly specific and restricted to the CD4-positive T cell population. T cells from pemphigus vulgaris patients with no anti-Dsg1 serum reactivity showed a proliferative response to Dsg3, but not to Dsg1. The Dsg1 fusion protein used in this study has minimal sequence homology with Dsg3. Thus, this study provides the first evidence that T cells from a subset of pemphigus vulgaris patients respond to both Dsg1 and Dsg3.

Amino Acid Sequence↗
PubMed1997

SEB is cytotoxic and alters EC barrier function through protein tyrosine phosphorylation in vitro.

We studied whether Staphylococcal enterotoxin B (SEB) has direct effects on endothelial cells (EC) in the absence of effector cells or their products. Bovine or human pulmonary artery EC were grown to confluence on filters mounted in chemotaxis chambers. Barrier function was assessed by placing [14C]bovine serum albumin in the chamber and sampling the lower well for 14C activity. SEB exposures induced a significant (P < 0.001) dose- and time-dependent increase in albumin flux across both bovine and human EC monolayers. Albumin flux was temperature dependent, and cycloheximide pretreatment of the monolayers did not block the SEB-induced increase in permeability. Preincubation of SEB with trypsin or anti-SEB antibody significantly (P < 0.0001) reduced the effect, whereas pretreatment with polymyxin B did not. SEB at > or = 10 micrograms/ml significantly (P < 0.03) increased EC injury as measured by 51Cr release in a dose- and time-dependent manner. Herbimycin and genistein, inhibitors of protein tyrosine kinases, each protected against SEB-induced cytotoxicity, barrier dysfunction, and intercellular gap formation. We conclude that SEB perturbs endothelial barrier function and viability in the absence of effector cells or their mediators.

Animals↗
PubMed1997

[Preparation of liposomes of superoxide dismutase].

The liposomes of SOD were prepared by reverse-phase evaporation. By examining the important effects on preparation we obtained the liposomes of SOD. The encapsulation of SOD was more than 60%, and the stability of the liposomes was excellent.

Delayed-Action Preparations↗
PubMed1997

[Studies on the effective constituents of arresting bleeding of Schizonepeta tenuifolia].

Schizonepeta tenuifolia (Benth.) Briq fried with carbon is used in various bleeding diseases in order to further investigate the effective constituents of arresting bleeding in Schizonepeta tenuifolia(benth.) Briq and to finish the quality standard of its capsule. We isolate the chemical constituents from Schizonepeta tenuifola(Benth.) Briq fried with carbon and study its action of arresting bledding and assay the effective constituents of arresting bleeding in capsule.

Animals↗
PubMed1997

[Analysis of triacylglycerol composition of perilla oil by combination of non-aqueous reversed-phase high performance liquid chromatography with gas chromatography].

Perilla oil was verified to be a rich source of polyunsaturated fatty acid (PUFA). Its linolenic content is 64.82%, the highest in the plants. Its triacylglycerol (TAG) components were isolated and identified first time in this paper by a combination of non-aqueous reversed-phase high performance liquid chromatography (RP-HPLC) with gas chromatography (GC). The TAG of perilla oil were isolated by HPLC with a Zorbax ODS column (5 microm, 4.6 mm i.d. x 250 mm; Dupont, Inc) and differential refractometer. The eluent was acetone/acetonitrile (80:20, V/V) with flow rate of 1.0 mL/min. The TAG component acyl groups were converted to fatty acid methyl esters (FAME). The acyl constituents for each TAG were determined by GC analysis. Gas chromatography of FAMEs was performed with a AC20 Carbowax 20M column (30 m x 0.32 mm i.d.). Detector and injection port temperatures were 260 degrees C. Column temperature was programmed from 120 degrees C, 1 min initially hold, then 8 degrees C/min to 220 degrees C and finally hold for 10 min. Nitrogen was the carrier gas. FAMEs peaks were identified by comparison of their retention times with those of standards. The retention times of the palmitic, stearic, oleic, linoleic, n-nonadecanoic acid (internal standard) and linolenic FAMEs were 10.7, 13.0, 13.3, 13.8, 14.2 and 14.7 min respectively. Five main components were determined. They were LnLnLn (34.10%), LnLnL (13.22%), LnLL (6.22%), LnLnO (11.38%) and LnLnP (9.17%). The TAG contents were corrected from their fatty acids content. The combination of non-aqueous RP-HPLC with GC is a simple and rapid method for the analysis of oil TAG structure and is also very helpful for the identification of oil quality.

Chromatography, Gas↗
PubMed1997

Nonsecretory IgA1 autoantibodies targeting desmosomal component desmoglein 3 in intraepidermal neutrophilic IgA dermatosis.

Intraepidermal neutrophilic IgA dermatosis, a rare skin disease entity manifested with blisters and pustules clinically and lower epidermal blister, acantholysis, and neutrophilic infiltration pathologically, was first reported in 1985. Although the disease is characterized by IgA autoantibodies targeting the epithelial cell surface component, the target antigen has not been determined. We investigated a patient with this disease by histopathology, direct and indirect immunofluorescence, immunoblotting, and immunoadsorption studies. The pustular lesion was characterized by blister at the lower epidermis, acantholysis, and neutrophilic infiltration. Nonsecretory IgA1 subclass autoantibodies targeting the lower epithelial cell surfaces were detected in the patient's skin and serum. The patient's IgA autoantibodies labeled a recombinant desmosomal protein desmoglein 3 on immunoblotting and the immunolabeling of epithelial cell surfaces was eliminated by preadsorption with desmoglein 3. Thus, desmoglein 3 is identified as a target antigen in intraepidermal neutrophilic IgA dermatosis. The ability of IgA1 autoantibodies to bind neutrophils may be responsible for the prominent neutrophilic infiltration observed histopathologically and for the pustular lesions observed clinically.

Aged↗
PubMed1997

Segmented three-dimensional echo-planar flow imaging of the cervical carotid arteries.

PURPOSE: To implement and assess the application of segmented three-dimensional echo-planar MR imaging time-of-flight flow sequences for studying the anatomy of the cervical carotid arteries at 1.5 T. METHODS: The 3-D echo-planar sequences were segmented along the in-plane phase-encoding direction. Echo train lengths (ETLs) of 3 and 5 and signal bandwidths of +/-25, +/-33, and +/-50 KHz were tested along with a conventional (ETL = 1) 3-D MR flow study in six healthy volunteers and in five patients with known arteriosclerotic disease involving the carotid bifurcation as confirmed by conventional angiography. The volunteer data were used to rank the techniques with respect to vessel dimension, vessel/background contrast, and quality by four trained neuroradiologists. For the patient studies, the percentage of stenoses was measured for all MR studies and compared against the conventional angiographic data using the criteria of the North American Symptomatic Carotid Endarterectomy Trial. RESULTS: Using Wilcoxon's test statistic and a significance level of .05, we found that the conventional MR flow examination was better than the segmented techniques and that the segmented techniques with ETL of 3 were superior to their counterparts with ETL of 5. For the ETL of 3 techniques, the high-bandwidth studies were inferior to their lower bandwidth counterparts; however, there was no significant difference between the performance of the medium- and low-bandwidth sequences. The patient data revealed that the segmented techniques consistently overestimated the severity of stenosis; however, in no instance did any of the segmented examinations erroneously indicate the presence of disease. CONCLUSIONS: The reduction in acquisition time and the zero false-positive rate we obtained suggest that segmented 3-D echo-planar MR flow techniques may be used as a screening/locating study for cervical carotid artery disease.

Adult↗
PubMed1997

Cellular mechanisms controlling protein degradation in catabolic states.

The daily turnover of protein amounts to 280 g in an adult weighing 70 kg but the metabolic processes responsible for protein turnover are only just beginning to be understood. In cells, the major pathway of protein degradation is the ubiquitin-proteasome pathway and protein flux through this pathway is precisely regulated. In catabolic conditions such as uremia, activity of the ubiquitin-proteasome pathway increases, resulting in degradation of muscle protein. In addition to increased protein degradation, gene transcription is activated, resulting in higher levels of the mRNAs encoding ubiquitin and proteasome subunits. The signals activating this pathway include metabolic acidosis and glucocorticoids but must be more diverse since the pathway is also activated in response to starvation, sepsis, cancer, muscle denervation, thermal injury, and acute diabetes. Understanding how the pathway is controlled could lead to the prevention of muscle loss in uremia and other conditions.

Animals↗