Biomedical subjects

L Wang

Publications and source records attributed to L Wang.

At least 829 records · Page 46Linked to original sources

PubMed2000

Relative abundance of placental pro-atrial natriuretic factor mRNA in normal pregnancy and pre-eclampsia.

Atrial natriuretic factor (ANF), produced by cytotrophoblast cells of the human placenta, may be involved in the regulation of uteroplacental blood flow. Pre-eclampsia is associated with maternal hypertension and reduced uteroplacental perfusion. The relationship between pre-eclampsia and placental production of ANF is not known. This study measured pro-ANF mRNA levels by Northern blot analysis in placentae delivered by caesarean section at preterm and term gestations from women with normotensive and pre-eclamptic pregnancies and found no significant difference between pre-eclampsia and normal pregnancy at either gestation. This result suggests that placental production of ANF is not altered at the pretranslational level during pre-eclampsia.

Adult↗

PubMed2000

Cardiovascular effects of 2-arachidonoyl glycerol in anesthetized mice.

Cannabinoids, including the endogenous ligand anandamide, elicit pronounced hypotension and bradycardia through the activation of CB1 cannabinoid receptors. A second endogenous cannabinoid, 2-arachidonoyl glycerol (2-AG), has been proposed to be the natural ligand of CB1 receptors. In the present study, we examined the effects of 2-AG on mean arterial pressure and heart rate in anesthetized mice and assessed the role of CB1 receptors through the use of selective cannabinoid receptor antagonists and CB1 receptor knockout (CB1(-/-)) mice. In control ICR mice, intravenous injections of 2-AG or its isomer 1-AG elicit dose-dependent hypotension and moderate tachycardia that are unaffected by the CB1 receptor antagonist SR141716A. The same dose of SR141716A (6 nmol/g IV) completely blocks the hypotensive effect and attenuates the bradycardic effect of anandamide. 2-AG elicits a similar hypotensive effect, resistant to blockade by either SR141716A or the CB2 antagonist SR144528, in both CB1(-/-) mice and their homozygous (CB1(+/+)) control littermates. In ICR mice, arachidonic acid (AA, 15 nmol/g IV) elicits hypotension and tachycardia, and indomethacin (14 nmol/g IV) inhibits the hypotensive effect of both AA and 2-AG. Synthetic 2-AG incubated with mouse blood is rapidly (<2 minutes) and completely degraded with the parallel appearance of AA, whereas anandamide is stable under the same conditions. A metabolically stable ether analogue of 2-AG causes prolonged hypotension and bradycardia in ICR mice, and both effects are completely blocked by SR141716A, whereas the same dose of 2-AG-ether does not influence blood pressure and heart rate in CB1(-/-) mice. These findings are interpreted to indicate that exogenous 2-AG is rapidly degraded in mouse blood, probably by a lipase, which masks its ability to interact with CB1 receptors. Although the observed cardiovascular effects of 2-AG probably are produced by an arachidonate metabolite through a noncannabinoid mechanism, the CB1 receptor-mediated cardiovascular effects of a stable analogue of 2-AG leaves open the possibility that endogenous 2-AG may elicit cardiovascular effects through CB1 receptors.

Anesthesia↗

PubMed2000

The role of the LDL receptor in apolipoprotein B secretion.

Familial hypercholesterolemia is caused by mutations in the LDL receptor gene (Ldlr). Elevated plasma LDL levels result from slower LDL catabolism and a paradoxical lipoprotein overproduction. We explored the relationship between the presence of the LDL receptor and lipoprotein secretion in hepatocytes from both wild-type and LDL receptor-deficient mice. Ldlr(-/-) hepatocytes secreted apoB100 at a 3.5-fold higher rate than did wild-type hepatocytes. ApoB mRNA abundance, initial apoB synthetic rate, and abundance of the microsomal triglyceride transfer protein 97-kDa subunit did not differ between wild-type and Ldlr(-/-) cells. Pulse-chase analysis and multicompartmental modeling revealed that in wild-type hepatocytes, approximately 55% of newly synthesized apoB100 was degraded. However, in Ldlr(-/-) cells, less than 20% of apoB was degraded. In wild-type hepatocytes, approximately equal amounts of LDL receptor-dependent apoB100 degradation occured via reuptake and presecretory mechanisms. Adenovirus-mediated overexpression of the LDL receptor in Ldlr(-/-) cells resulted in degradation of approximately 90% of newly synthesized apoB100. These studies show that the LDL receptor alters the proportion of apoB that escapes co- or post-translational presecretory degradation and mediates the reuptake of newly secreted apoB-containing lipoprotein particles.

Adenoviridae↗

PubMed2000

Factors associated with vagal tone responses in preterm infants.

The purpose of this study was to examine factors related to vagal tone (VNA) among preterm infants receiving a 10-minute gentle human touch (GHT) intervention three times daily for 10 days. VNA was measured continuously for 10 minutes before, during, and after each 10-minute GHT intervention. Findings indicated that there was a significant relationship between VNA and gestational age, although there were no relationships between VNA and measures of motor activity or behavioral distress. There was no difference in pattern of response to GHT or level of morbidity, average daily weight gain, or behavioral organization among infants with low, moderate, and high baseline VNA levels. There was no difference in VNA comparing infants in the GHT and control groups or during baseline, touch, and posttouch phases for infants in the GHT group. There is a need for further research to examine the usefulness of VNA as a measure of stress vulnerability among preterm infants.

Female↗

PubMed2000

Stage-specific expression of pituitary adenylate cyclase-activating polypeptide type I receptor messenger ribonucleic acid during ovarian follicle development in the rat.

Expression of pituitary adenylate cyclase-activating polypeptide (PACAP), a neuropeptide with considerable homology to vasoactive intestinal peptide, has been shown to be stimulated by gonadotropins in the ovary. The present studies further evaluated the cell-type specific expression and gonadotropin regulation of PACAP type I receptor (PACAPR) messenger RNA in immature rat ovaries and in cultured preovulatory follicles. Northern blot analysis of ovaries obtained from prepubertal rats revealed the increased expression of PACAPR during prepubertal development. The major cell types expressing PACAPR messenger RNA were granulosa cells of large preantral follicles. Treatment of immature rats with PMSG caused a decrease in ovarian PACAPR expression. In contrast, treatment with human (h) CG at 2 days after PMSG treatment stimulated ovarian PACAPR messenger RNA within 3-6 h in granulosa cells of preovulatory follicles. Treatment of cultured preovulatory follicles in vitro with LH further confirmed the time- and dose-dependent stimulation of PACAPR by gonadotropins in granulosa cells of preovulatory follicles. Moreover, RNase protection assay revealed that the short variant of ovarian PACAPR was the predominant form stimulated during prepubertal development and by gonadotropins. These results demonstrate the expression of PACAPR messenger RNA in granulosa cells of growing follicles and of preovulatory follicles stimulated by gonadotropins, and suggest that PACAP may play a role in the growth of developing follicles and in ovulation as an autocrine/paracrine factor.

Aging↗

PubMed2000

Two putative GATA motifs in the proximal exon 1 promoter of the rat insulin-like growth factor I gene regulate basal promoter activity.

The insulin-like growth factor I gene is transcribed from two promoters, which direct synthesis of alternative first exons (exon 1 and exon 2) in insulin-like growth factor I messenger RNAs (mRNAs). An exon 1 promoter construct extending from +75 to +192 (the most upstream exon 1 start site was designated as +1) showed significant promoter activity in C6, OVCAR-3, and SK-N-MC cells. Within the +75 to +192 region, there are two perfect matches to the consensus binding site for GATA transcription factor, at +108 (GATA-A) and at +183 (GATA-B). Mutations of the GATA-A or GATA-B sequences resulted in slight (or no) effect on exon 1 promoter activity in both C6 and OVCAR-3 cells. However, mutation of the GATA-A sequence stimulated exon 1 promoter activity by 68% in SK-N-MC cells. Mutation of the GATA-B sequence inhibited exon 1 promoter activity by 4.4-fold in SK-N-MC cells. Electrophoretic mobility shift assays showed that there were nuclear proteins in SK-N-MC cells capable of specifically binding to the GATA-A and GATA-B elements and that this binding was GATA-sequence specific. GATA-2, GATA-3, and GATA-4 are the only GATA proteins that have been reported to be expressed in neurons. None of the antibodies against these three GATA proteins were capable of inhibiting or supershifting the bands formed by the nuclear proteins and oligonucleotides containing GATA-A or GATA-B elements. A GATA-1 expression vector was used to perform cotransfection experiments. The GATA-A mutation abolished the stimulatory effect of the GATA-1 factor on promoter activity. In contrast, the GATA-B mutation enhanced the stimulatory effect of GATA-1 protein. Anti-GATA-1 antibody was also incapable of inhibiting or supershifting the bands formed by the nuclear proteins and oligonucleotides containing the GATA-A or GATA-B elements. In conclusion, the GATA-A element seems to bind an inhibitory endogenous factor(s) in SK-N-MC cells, whereas the GATA-B element may bind a stimulatory factor(s). These factors seem to be related to GATA transcription factors but are immunologically distinct from GATA-2, GATA-3, or GATA-4. GATA-1 has the potential to transactivate the exon 1 promoter through the GATA-A element but is unlikely to be the endogenous protein binding to the GATA-A or the GATA-B motifs in SK-N-MC cells.

Animals↗

PubMed2000

Cell density influences insulin-like growth factor I gene expression in a cell type-specific manner.

The effect of cellular density on insulin-like growth factor I (IGF-I) gene expression was characterized in several tumor-derived cell lines. IGF-I messenger RNA (mRNA) transcripts increased more than 200-fold when C6 glioma cells grew to postconfluence. IGF-I receptor and beta-actin mRNAs were induced by 6- and 2-fold, respectively, as a function of confluence. IGF-I mRNA transcripts in GH3 and SK-N-MC cells increased about 4- to 5-fold in confluent cultures compared with sparse cultures. In OVCAR-3 cells, the IGF-I mRNA level remained constant as the cell density increased. Transient transfection experiments were performed with IGF-I exon 1 promoter/luciferase fusion constructs in C6 cells. The luciferase activity of a construct containing exon 1 sequence between +75 and +282 (the most 5' transcription initiation site was designated +1) was stimulated by 2.5-fold in dense cultures compared with that in sparse cultures of C6 cells. Luciferase activities of other constructs containing at least 282 bp of exon 1 sequence were also stimulated about 2- to 4-fold by cell density. However, 3' deletion to +192 led to loss of the cell density stimulatory effect. In contrast, luciferase activities of IGF-I promoter constructs were not altered by cell density in SK-N-MC cells. When the conditioned medium of low density C6 cultures was exchanged with that of high density cultures, the IGF-I mRNA level remained the same. In summary, cell density has a cell type- and gene type-specific effect on IGF-I gene expression. A cell density response element(s) may be located between +192 and +282 of the exon 1 promoter region in C6 cells.

Actins↗

PubMed2000

Comparison of alendronate and intranasal calcitonin for treatment of osteoporosis in postmenopausal women.

This study compared the effects of oral alendronate and intranasal calcitonin for treatment of osteoporosis in postmenopausal women. Women at least 5 yr postmenopause (n = 299) were randomized to either 10 mg alendronate, matching alendronate placebo, or open-label intranasal calcitonin 200 IU daily for 12 months. Hip and spine bone mineral density (BMD) and markers of bone turnover were measured, and safety and tolerability were assessed. Alendronate produced greater increases in BMD than calcitonin at 12 months at the lumbar spine (5.16% vs. 1.18%; P < 0.001), trochanter (4.73% vs. 0.47%; P < 0.001), and femoral neck (2.78% vs. 0.58%; P < 0.001). Changes in BMD with calcitonin were greater than with placebo at the femoral neck, but were not different from placebo at either the trochanter or lumbar spine. Greater decreases in bone turnover were seen with alendronate than with calcitonin (serum bone-specific alkaline phosphatase, 43% vs. 9%, P < 0.001; urinary N-telopeptide, 62% vs. 11%, P < 0.001). Similar percentages of patients in each group reported an adverse experience during the study. We conclude that, in postmenopausal women with osteoporosis, 12 months of therapy with alendronate produced significantly greater increases in BMD of the hip and spine and greater decreases in bone turnover than intranasal calcitonin.

Absorptiometry, Photon↗

PubMed2000

Effects of the Chinese medicine, TSJN on insulin resistance and hypertension in fructose-fed rats.

The aim of this study was to determine the effect of Tang-Shen-Jiao-Nang (TSJN), a Chinese medicine used to treat diabetes mellitus, on insulin resistance and hypertension in fructose-fed rats (FFR). Six-week-old male Sprague-Dawley rats were fed either normal rat chow (control) or a fructose-rich chow (FFR) for 6 wk. For the last 2 or 4 wk of a 6-wk period of either diet, the rats were treated by gavage with gum arabic solution as a vehicle (control or FFR) or TSJN (800 mg/kg/d; FFR+TS), and then we performed the euglycemic hyperinsulinemic glucose clamp technique to estimate insulin sensitivity. Systolic blood pressure was measured weekly for 6 wk. At the end of the glucose clamp, the soleus muscle was dissected out for determination of muscle fiber composition by ATPase methods. Systolic blood pressure was elevated at 2 wk after the start of the fructose-rich chow feeding and persisted thereafter throughout the study. Systolic blood pressure during the glucose clamp in the FFR group was significantly higher than that in the control group. Although there was no effect on systolic blood pressure in rats treated with TSJN for the last 2 wk of their 6-wk diet, those treated with TSJN for the last 4 wk of their 6-wk diet had lower systolic blood pressure than did the rats in the FFR group. The average rate of glucose infusion during the glucose clamp, as a measure of insulin sensitivity (M value), was significantly lower in the FFR than in the controls (10.9 +/- 0.6 and 15.4 +/- 0.4, mg/kg/min, for FFR and controls, respectively; p< 0.01). Treatment with TSJN for 2 wk significantly improved the M value compared to that of the control level (15.1 +/- 0.5 mg/kg/min). The composite ratio of type I fibers in the soleus muscle was significantly decreased in the FFR compared to controls (75.0 +/- 1.7 and 81.7 +/- 1.5%, for FFR and controls, respectively; p< 0.01), and treatment with TSJN for 2 wk led to a recovery composite ratio of type I fiber to the same level as that of the control group (78.7 +/- 1.7% in FFR + TS). The M value was significantly correlated with the compositions of type I and type II fibers (for type I fibers, r= 0.45, p < 0.01, for type II fibers, r= -0.44, p< 0.05). These results suggest that the Chinese medicine TSJN may improve insulin resistance, lower the systolic blood pressure, and modulate muscle fiber composition in hypertensive and insulin-resistant fructose-fed rats.

Animals↗

PubMed2000

Ionizing radiation-induced mitogen-activated protein (MAP) kinase activation in DU145 prostate carcinoma cells: MAP kinase inhibition enhances radiation-induced cell killing and G2/M-phase arrest.

These studies examine the role(s) played by the mitogen-activated protein kinase (MAPK) pathway after exposure of DU145 prostate carcinoma cells to radiation. Radiation (2 Gy) was found to cause both immediate primary (0-30 min) and prolonged secondary activations (90-1440 min) of the MAPK pathway. These activations of the MAPK pathway were abolished by inhibition of epidermal growth factor receptor (EGFR) function. The secondary activation was also abolished by addition of a neutralizing monoclonal antibody against transforming growth factor alpha (TGFA). Activation of the MAPK pathway could be induced in nonirradiated cells by the transfer of medium from irradiated cultures. Neutralizing antibody to TGFA blocked this effect, indicating that radiation causes secondary activation of the MAPK pathway by release of TGFA in DU145 cells. Radiation induced a transient G(2)/M-phase growth arrest that was prolonged for up to 24 h by inhibition of the MAPK pathway. Inhibition of the MAPK pathway significantly increased the ability of radiation to cause apoptosis 24 h after exposure. The ability of DU145 cells to proliferate after irradiation became dependent on MAPK signaling. When cells were subjected to single doses or fractionated radiation exposure, continuous inhibition of the MAPK pathway significantly decreased clonogenic survival. Only a small fraction of this cell killing could be accounted for by apoptosis within the first 96 h. Thus inhibition of the MAPK pathway increased radiation-induced cell killing likely by both apoptotic and nonapoptotic mechanisms. Collectively, our findings indicate that disruption of the TGFA/EGFR/MAPK pathway may represent a strategy that could be exploited to manipulate prostate carcinoma growth and cell survival after irradiation.

Apoptosis↗

PubMed2000

Immediate post-abortal insertion of the frameless GyneFix IUD: initial experience in China.

The promising results obtained with the frameless GyneFix IUD for interval insertion led to the belief that the technology could be useful for immediate post-abortal application to reduce the number of induced and repeat abortions. IUDs have many advantages and the method is acceptable and safe. However, side effects and expulsion of conventional IUDs remain a problem. In an attempt to minimise these problems, the frameless GyneFix IUD was developed. Clinical studies conducted over the past 14 years have shown that the principles on which the device is based are valid. An increasing bulk of clinical evidence shows that the design characteristics of the device (fixed, frameless and flexible) are responsible for the low expulsion, high effectiveness, and high tolerance and continuation rates.1 Experience with the frameless IUD in China since 1989 confirms the international experience with the device. The present communication is a second report from China with the GyneFix IUD. In the present study the GyneFix IUD is inserted immediately following termination of pregnancy of less than 10 weeks amenorrhea. It is concluded that immediate post-abortal insertion of the frameless IUD is easy and safe, and appears to be as reliable and as effective as when it is inserted at interval. The GyneFix IUD could, therefore, constitute an important new option in the prevention of abortion.

Abortion, Induced↗

PubMed2000

Cloning of a T-type Ca2+ channel isoform in insulin-secreting cells.

The T-type Ca2+ channel is an important determinant of electrical activity and of Ca2+ influx in rat and human pancreatic beta-cells. We have identified and sequenced a cDNA encoding a T-type Ca2+ channel alpha1-subunit derived from INS-1, the rat insulin-secreting cell line. The sequence of the cDNA indicates a protein composed of 2,288 amino acids that shares 96.3% identity to alpha1G, the neuronal T-type Ca2+ channel subunit. The transmembrane domains of the protein are highly conserved, but the isoform contains three distinct regions and 10 single amino acid substitutions in other regions. Sequencing rat genomic DNA revealed that the alpha1-subunit we cloned is an alternative splice isoform of alpha1G. By using specific primers and reverse transcription-polymerase chain reaction, we demonstrated that both splice variants are expressed in rat islets. The isoform deduced from INS-1 was also expressed in brain, neonatal heart, and kidney. Functional expression of this alpha1G isoform in Xenopus oocytes generated low voltage-activated Ba2+ currents. These results provide the molecular biological basis for studies of function of T-type Ca2+ channels in beta-cells, which is where these channels may play critical roles in diabetes.

Alternative Splicing↗

PubMed2000

Factors affecting hepatocyte viability and CYPIA1 activity during encapsulation.

Hepatocytes encapsulated in alginate-poly-1-lysine-alginate (APA) are used in transplantation studies and in bioartificial liver support systems. Loss of cell viability in the process of APA encapsulation is usually 20-30% while the effect on cytochrome CYP450 activity is rarely reported. This work investigates the negative influences on hepatocyte viability and CYPIA1 activity during APA encapsulation, and reports methods to alleviate these influences by incorporating certain reagents into the encapsulation solution. The results show that loss of hepatocyte viability and CYPIA1 activity was caused almost entirely by extracellular calcium toxicity rather than by mechanical damage (p < 0.05). Use of 10 mM instead of 100 mM calcium chloride (CaCl2) in the encapsulation process improved CYPIA1 activity (p < 0.05), but did not improve hepatocyte viability (p > 0.05) or result in satisfactory microcapsules. Hepatocyte viability was 25% higher (p < 0.05) in CaCl2 than in calcium lactate (CaLa) when the cells were gelled by contact with these calcium solutions at room temperature (RT). Hepatocyte viability showed little improvement by processing at 4 degrees C than at RT in CaCl2 (p > 0.05) but was 23% higher at 4 degrees C than at RT in CaLa (p < 0.05). Calcium used in the process of encapsulation caused cell necrosis rather than apoptosis. Addition of Dulbecco's modified Eagle's medium (containing 10% foetal bovine serum) or 20 mM fructose to the calcium solution did not improve cell survival. However, nifedipine at a final concentration of 25 mM modestly improved hepatocyte survival in solution containing 100 mM CaCl2 (p = 0.003). Glutathione and taurine in certain concentrations showed protective effects against loss of CYPIA1 activity (p < 0.05 and <0.01 respectively). In conclusion, to optimise the use of calcium during the process of encapsulation, CaCl2 is preferred to CaLa and inclusion of nifedipine, glutathione or taurine in 100 mM CaCl2 solution is recommended.

Alginates↗

PubMed2000

Expression of SAA and amyloidogenesis in congenic mice of CE/J and C57BL/6 strains.

Inbred strains of mice display different susceptibilities to experimental induction of reactive amyloidosis. CBA/J, C57BL/6, and ICR are among the most susceptible strains, while CE/J mice appear to be totally resistant. In contrast to amyloidogenic strains which express two major acute phase serum amyloid A proteins (SAA1 and SAA2), CE/J mice produce only a single isoform, designated SAA2.2. Studies indicate that CE/J x CBA/J hybrid mice expressing both SAA2.2 and SAA1.1/SAA2.1 are amyloid resistant, and this has led to the hypothesis that SAA2.2 may protect mice against amyloid formation even in the presence of fibrillogenic SAA1.1. We have tested this hypothesis in mice derived from CE/J and C57BL/6 strains. CE/J mice were mated with C57BL/6 mice to produce F1 hybrids. Congenic mice were then produced by backcrossing each successive generation to C57BL/6 mice. Representative mice from F2 and F3 generations were analyzed to determine SAA genotype and susceptibility to amyloid induction by repeated casein injections. All F2 and F3 mice examined, including those which carried the SAA2.2 gene, developed extensive splenic AA amyloid. Expression of SAA2.2 in mice testing positive for the SAA2.2 gene was confirmed by sequence analysis of HDL-associated SAA proteins. These results demonstrate that the presence of SAA2.2 is not sufficient to protect CE/J x C57BL/6 hybrid mice from amyloid development. This is consistent with our observation that macrophage cultures, derived from C57BL/6, CBA/J, or CE/J mice, undergo amyloid deposition when treated with SAA1.1 alone or with equal amounts of SAA1.1 and SAA2.2, but show no deposition when treated solely with SAA2.2. We conclude from these studies that while SAA2.2 is non-fibrillogenic, its physical presence is not sufficient for protection against amyloid formation.

Amyloidosis↗

PubMed2000

Menopausal symptoms: experience of Chinese women.

OBJECTIVES: To investigate the prevalence of symptoms in middle-aged Chinese rural and city women of different occupations; and to explore the relationship between symptoms, hormone levels and other factors. METHODS: This cross-sectional study included 806 women aged 41-60 years, selected by multistage cluster sampling and a structured interview questionnaire. The response rate was 95%. The sample compromised 402 professional urban women and 404 women farmers living in rural areas. Some 209 women were randomly selected from the two groups for hormonal assay and bone mineral density screening. RESULTS: The professional group was more symptomatic than the farming group (p < 0.01). The presence of symptoms was significantly related to an increasing level of education. There were no significant differences between occupational groups in levels of estradiol follicle stimulating hormone (FSH) and luteinizing hormone (LH). Estradiol level and bone mineral density were decreased and FSH and LH increased in the postmenopausal group (p < 0.05) compared with the other menopausal groups. Hot flushes were the only symptom to be significantly associated with hormone levels (estradiol and LH). The occurrence of symptoms was significantly related to indications of bone and joint disease, heart disease, primary dysmenorrhea, decline in sexual interest, irregular menses and feelings of becoming older, sad and lost. These factors were all reported more often by the professional women than by the farmers. CONCLUSION: Symptom experience in mid-life Chinese women is related to both biological and psychosocial factors.

Adult↗

PubMed2000

Cellular origin of gastrointestinal stromal tumors: a study of 27 cases.

CONTEXT: Interstitial cells of Cajal (ICCs), also known as pacemaker cells, are cells in the gastrointestinal tract that play a role in the control of gut motility. The ICCs express the c-kit proto-oncogene encoding a type III tyrosine kinase (KIT) receptor, a ligand that is known as stem cell factor (SCF). The maturation of ICCs is dependent on SCF-KIT interaction. The cellular origin, differentiation, nomenclature, and prognosis of gastrointestinal stromal tumors (GISTs) are controversial. OBJECTIVE: To test the hypothesis that GISTs originate from CD34-positive stem cells and differentiate toward an ICC phenotype. MATERIALS AND METHODS: We studied 27 cases of smooth muscle differentiated GISTs collected for 14 years (1985-1999), including 8 benign (leiomyoma), 15 malignant primary (leiomyosarcoma), and 4 metastatic to the liver. Immunohistochemical studies of selected lineage-directed monoclonal antibodies of c-kit (CD117), CD34, vimentin, desmin, alpha-actin, S100, and MIB-1 were performed on both normal and tumor tissues. RESULTS: Immunoperoxidase stains of normal gastrointestinal tract showed both c-kit and CD34-positive cells surrounding the Auerbach ganglia plexus in the gastrointestinal tract. Twenty-seven of 27 tumors strongly expressed c-kit. Fourteen of 27 tumors were positive for CD34. Of the malignant GISTs, 14 of 19 were positive for CD34; of the benign tumors, 0 of 8 were positive for CD34. Thus, CD34 was the best indicator of malignant phenotype. CONCLUSION: This is the first description of benign smooth muscle GISTs negative for CD34. The results of this study suggest that GISTs originate from CD34-positive stem cells and differentiate toward pacemaker cell phenotype. The lack of expression of CD34 in the benign GIST may indicate that benign GISTs are composed of more mature ICCs, whereas malignant GISTs are composed of dedifferentiated ICCs that express CD34-positive stem cells.

Adult↗

PubMed2000

[A convenient method of recovering DNA from agarose electrophoretic gels].

OBJECTIVE: Construct a simple unit to efficiently recover DNA from agarose electrophoretic gels. METHODS: With the use of 1.5ml microcentrifuge tube, 1 ml tip, nylon web membrane and dialysis membrane, the authors made a clever yet simple DNA recovering gadget, tested this gadget by using it to recover specific electrophoretic DNA bands of lambdaDNA/EcoR I+Hind III markers, and then examined the recovery ratio through agarose gel electrophoresis. A restriction enzyme's digestion product was taken as another sample to test the recovered product's quality by means of DNA ligation and subsequent agarose gel electrophoresis. RESULTS: Under such electroelution conditions as with an electrophoretic buffer of 1xTBE, a voltage grade of 3V/cm and an elution time of 0.5h, the final recovery ratio of 564bp DNA was up to 80%, and that of 21227bp DNA was about 30%. Judged either by the agarose gel electrophoresis performance or by the DNA ligation result, the recovered DNA proved to be of excellent quality. CONCLUSION: It is an easy, quick, efficient and cheap method of recovering DNA from agarose electrophoretic gels. The recovering unit can be a permanent one, provided that the dialysis membrane in this unit is changed for recovering different DNA samples.

DNA↗

PubMed2000

Neurotensin receptor-1 mRNA analysis in normal pancreas and pancreatic disease.

By autoradiography, neurotensin (NT) binding is specifically detectable in pancreatic cancer, but not in the normal pancreas, chronic pancreatitis (CP), or other pancreatic disorders. In the present study, we investigated whether this is due to NT receptor-1 (NTR-1) mRNA up-regulation and whether NTR-1 mRNA could also be used as a specific diagnostic marker and treatment target in pancreatic cancer. Fifteen normal pancreas tissue samples, 20 CP samples, and 30 pancreatic cancer samples were studied. Expression and localization of NTR-1 mRNA was investigated by Northern blot analysis and in situ hybridization. Furthermore, consecutive tissue sections were analyzed for NTR-1 mRNA expression and NT binding. By Northern blot analysis, NTR-1 mRNA expression was 4.4-fold (P < 0.01) and 3.0-fold (P < 0.01) higher in pancreatic cancer and CP tissue samples, respectively, compared with normal controls. There was no difference in NTR-1 mRNA levels between CP and cancer samples (P > 0.05). In pancreatic cancer, the NTR-1 mRNA levels were higher in advanced tumor stage (stages III and IV) than early tumor stage (stages I and II; P < 0.05), but no difference was found between well/moderately differentiated (grades 1 and 2) and poorly differentiated/undifferentiated cancers (grades 3 and 4; P > 0.05). By in situ hybridization, NTR-1 mRNA signals were weakly present in the cytoplasm of acinar and ductal cells of the normal pancreas. Moderate to intense NTR-1 mRNA signals were present in the cytoplasm of acinar cells dedifferentiating into tubular complexes and degenerating acinar cells of CP samples. In the cancer samples, NTR-1 mRNA was moderately to intensely expressed in the cytoplasm of cancer cells. When on consecutive tissue sections NTR-1 mRNA expression was compared with the presence of NTR-1, measured by receptor autoradiography, a correlation was found in carcinomas but not in CP samples, in which no receptors were detectable by autoradiography. The enhanced expression of NTR-1 mRNA in pancreatic cancer cells further suggests that neuroendocrine hormones might modulate pancreas cancer cell behavior. However, its relatively high levels in CP excludes NTR-1 mRNA as a specific parameter for pancreatic cancer and for the differentiation of pancreatic cancer from CP.

Adult↗