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Biomedical subjects

S D Smith

Publications and source records attributed to S D Smith.

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

A comparison of neonatal and adult multiorgan failure in a rat model.

BACKGROUND/PURPOSE: To establish a neonatal animal model of multiorgan failure (MOF) for the histological study of the sequence and severity of neonatal MOF in comparison to a model of adult MOF. METHODS: Neonatal and adult Sprague-Dawley rats received a single intraperitoneal injection of the inflammatory agent Zymosan. Rats were weighed; randomly killed on days 1 through 6; and heart, lung, liver, kidney, spleen, and ileocecum harvested for histological examination. RESULTS: Neonatal animals receiving Zymosan showed a significant increase in total body weight not seen in adults. The sequence and severity of MOF-induced organ damage were strikingly different in adult and neonatal animals. Mild lung damage was seen as early as day 1 in adult rats receiving Zymosan. This progressed to moderate damage by day 2 and severe damage by day 6. Lungs of neonatal rats receiving Zymosan showed only mild damage by day 4, which had progressed no further by day 6. Mortality rate was not significantly different between adult and neonatal animals receiving Zymosan. CONCLUSIONS: Zymosan can be used in a neonatal animal model to incite MOF. In the neonatal animal model of MOF there is (1) substantial early capillary leak as shown by increased body weight; (2) a unique progression of organ involvement-liver, kidney, lung compared with adult animals with MOF-lung, liver, kidney; and (3) relative sparing of the lung from injury. These findings are consistent with previous clinical observations of a difference in neonatal and adult MOF.

Age Factors↗
PubMed2000

Expression of endothelin receptor subtypes and their messenger RNAs in diabetic rat prostate: effect of insulin treatment.

Streptozotocin (STZ)-induced diabetes causes an upregulation in the expression of endothelin (ET) receptors in the rat prostate (Eur J Pharmacol 310:197, 1996). We examined the effects of insulin treatment, started 8 weeks after the induction of diabetes, on the expression and distribution of ET receptors and their respective mRNAs in the rat prostate. The densities, pharmacological properties and distribution of ET receptors in the rat prostate were examined using radioligand receptor binding and autoradiographic studies, and gene expression of ET receptors was evaluated utilizing the reverse transcription-polymerase chain reaction (RT-PCR). STZ-injected rats had smaller prostates and reduced serum testosterone levels than control and insulin treated diabetic animals. ET receptor density was shown to be significantly higher in the prostate from diabetic rats than those from either control or insulin treated diabetic animals. The pharmacological profile of prostatic ET receptors was similar in all groups (approximately 80% ET(A); 20% ET(B) subtype). ET receptors were predominantly localized to the prostatic stroma. Induction of diabetes increased the expression of mRNA levels of ET(A) and ET receptors, and insulin treatment reversed this upregulation to control levels. These results indicate that (1) ET receptor subtypes are expressed in the rat prostate as transcription and translation products; (2) insulin can normalize the diabetes-induced upregulation in the expression of ET receptors and their respective mRNAs; and (3) diabetes-induced regression of the prostate may involve an alteration in ET receptors.

Animals↗
PubMed2000

Temporal changes of cytokines and nitric oxide products in urine from renal transplant patients.

BACKGROUND: Acute rejection and urinary tract infection (UTI) both increase nitric oxide synthase (NOS) activity in urine from renal transplant patients. Also, with rejection, a regulatory interplay between nitric oxide (NO) and cytokines has been suggested. Thus, measurement of the temporal changes of NOS products and cytokines in urine will provide a strategy for the diagnosis of acute rejection and for its differentiation from UTI. METHODS: Soluble interleukins (ILs) and NOS-related products, cyclic GMP (cGMP), nitrate, and nitrite were measured in 192 urine samples consecutively collected from 13 patients within the first three months of transplantation. Sixty-seven additional urine specimens were collected randomly from 24 patients for follow-up analysis of the nitrate test. RESULTS: Among patients who experienced rejection, the percentage (%) binding of IL-2 increased within the first five days (P = 0.0004) after transplantation and one to five days prior to the clinical diagnosis (dx) of rejection (P = 0.02). Tumor necrosis factor-alpha, IL-6, and IL-8 increased at the time of rejection dx (P < or = 0.01). With UTI, IL-2 (P = 0.01) decreased one to five days prior to dx, and IL-10 (P = 0.003) increased one to five days after dx. Although cGMP and nitrate are dependent variables, cGMP increased (P < or =0.0009) with both rejection and UTI, and nitrate increased (P = 0.0001) with rejection and decreased (P = 0.0001) with UTI. Prior to formal dx (1 to 5 days), urine nitrate clearly differentiated rejection (3004 to 7451 micromol/L) from UTI (90 to 885 micromol/L) and controls (1059 to 3235 micromol/L). The additional 67 urines demonstrated that the sensitivity of the nitrate test for rejection and UTI was 100%. CONCLUSIONS: In renal transplant patients, specific temporal changes in urine cytokine levels do occur with acute rejection and UTI, but urine nitrate levels are the most precise at differentiating rejection from UTI.

Acute Disease↗
PubMed2000

Expression of BCL-2, BAX and BAK in the trophoblast layer of the term human placenta: a unique model of apoptosis within a syncytium.

The regulation of apoptosis in the syncytiotrophoblast is of particular interest because this is the only true syncytial epithelium in human cell biology. Nuclei characteristic of apoptotic cells have been localized to this syncytium especially in association with fibrin-containing fibrinoid deposits. The factors responsible for regulating cell death-like features in the trophoblast syncytium are unknown. We tested the hypothesis that fibrin was required for trophoblast apoptosis. TUNEL (terminal deoxynucleotidyltransferase-mediated dUTP end-labelling) staining to detect DNA fragmentation typical of apoptosis was performed in term human placentae revealing labelled nuclei associated with fibrin-type fibrinoid, as well as labelled nuclei in discrete areas of syncytiotrophoblast without fibrin. We also hypothesized that members of the BCL-2 family of apoptosis-associated proteins contribute to the regulation of syncytiotrophoblast apoptosis. To identify members of this protein family that might regulate trophoblast apoptosis, we assessed expression of three important members of the bcl-2 gene family. We used immunohistochemistry with monoclonal antisera against human BCL-2 and polyclonal antisera against human BAX and BAK to study paraffin-embedded sections of human term placentae (n=5) from uncomplicated pregnancies. The anti-apoptotic BCL-2 protein was expressed throughout the syncytium of normal villi with much less staining in cytotrophoblast. Staining was also seen adjacent to fibrin deposits and in syncytium overlying fibrin deposits. Expression of the pro-apoptotic BAX protein was undetectable in the syncytiotrophoblast, was expressed in rare cytotrophoblast and was prominent in connective tissue and perivascular cells within the villous core. Localization of a second pro-apoptotic protein, BAK, revealed immunoreactivity in isolated areas of intact syncytium of normal villi. Additionally, fibrin deposits were associated with intense BAK staining in both syncytiotrophoblast and cytotrophoblast. From these data, we speculate that modulation of BAK expression is one factor regulating apoptosis in human trophoblast.

Adult↗
PubMed2000

Tietz syndrome (hypopigmentation/deafness) caused by mutation of MITF.

Patients with Tietz syndrome have congenital profound deafness and generalised hypopigmentation, inherited in a fully penetrant autosomal dominant fashion. The pigmentary features and complete penetrance make this syndrome distinct among syndromes with pigmentary anomalies and deafness, which characteristically have patchy depigmentation and variable penetrance. Only one family has been reported with the exact features described in the original report of this syndrome. This family was reascertained and a missense mutation was found in the basic region of the MITF gene in family members with Tietz syndrome. Mutations in other regions of this gene have been found to produce Waardenburg syndrome type 2 (WS2), which also includes pigmentary changes and hearing loss, but in contrast to Tietz syndrome, depigmentation is patchy and hearing loss is variable in WS2.

Adolescent↗
PubMed2000

Apoptosis in human cultured trophoblasts is enhanced by hypoxia and diminished by epidermal growth factor.

Preeclampsia and fetal growth restriction are associated with placental hypoperfusion and villous hypoxia. The villous response to this environment includes diminished trophoblast differentiation and enhanced apoptosis. We tested the hypothesis that hypoxia induces apoptosis in cultured trophoblasts, and that epidermal growth factor (EGF), an enhancer of trophoblast differentiation, diminishes hypoxia-induced apoptosis. Trophoblasts isolated from placentas of term-uncomplicated human pregnancies were cultured up to 72 h in standard (PO(2) = 120 mm Hg) or hypoxic (PO(2) <15 mm Hg) conditions. Exposure to hypoxia for 24 h markedly enhanced trophoblast apoptosis as determined by DNA laddering, internucleosomal in situ DNA fragmentation, and histomorphology, as well as by the reversibility of the apoptotic process with a caspase inhibitor. Apoptosis was accompanied by increased expression of p53 and Bax and decreased expression of Bcl-2. Addition of EGF to cultured trophoblasts or exposure of more differentiated trophoblasts to hypoxia significantly lowered the level of apoptosis. We conclude that hypoxia enhances apoptosis in cultured trophoblasts by a mechanism that involves an increase in p53 and Bax expression. EGF and enhancement of cell differentiation protect against hypoxic-induced apoptosis.

Apoptosis↗
PubMed2000

Peroxisome proliferator-activated receptor-gamma modulates differentiation of human trophoblast in a ligand-specific manner.

The ligand-dependent nuclear receptor peroxisome proliferator-activated receptor-gamma (PPARgamma) regulates the differentiation of several tissues and cell types. PPARgamma was recently determined to be essential for murine placental development and differentiation. We therefore assessed the influence of PPARgamma on differentiation of human placental trophoblasts. We initially used immunohistochemistry to examine term human placentas for PPARgamma expression and found that PPARgamma is present in syncytiotrophoblasts and cytotrophoblasts in placental villi. We correlated the expression of PPARgamma with differentiation of primary human trophoblasts and found that 8-bromo-cAMP, a known enhancer of trophoblast differentiation, stimulates PPARgamma activity, but has no effect on PPARgamma expression. We demonstrated that the PPARgamma ligand 15-deoxy-delta12,14-prostaglandin J2 (15deltaPGJ2) and the thiazolidinedione troglitazone stimulate PPARgamma activity in the trophoblast cell line BeWo. Importantly, whereas exposure of cultured primary trophoblasts to troglitazone enhances biochemical and morphological trophoblast differentiation, 15deltaPGJ2 diminishes trophoblast differentiation. Furthermore, 15deltaPGJ2, but not troglitazone, up-regulates p53 expression and promotes trophoblast apoptosis. These data indicate that PPARgamma is expressed in human placental trophoblasts, and that ligand-specific activation of PPARgamma results in opposing effects on trophoblast differentiation. Our results suggest that PPARgamma plays an important role in placental differentiation during human pregnancy.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1999

Human connexin 30 (GJB6), a candidate gene for nonsyndromic hearing loss: molecular cloning, tissue-specific expression, and assignment to chromosome 13q12.

Mutations in connexin 26 are responsible for approximately 20% of genetic hearing loss and 10% of all childhood hearing loss. However, only about 75% of the mutations predicted to be in Cx26 are actually observed. While this may be due to mutations in noncoding regulatory regions, an alternative hypothesis is that some cases may be due to mutations in another gene immediately adjacent to Cx26. Another gap junction gene, connexin 30 (HGMW-approved symbol GJB6), is found to lie on the same PAC clone that hybridizes to chromosome 13q12. Human connexin 26 and connexin 30 are expressed in the same cells of the cochlea. Cx26 and Cx30 share 77% identity in amino acid sequence but Cx30 has an additional 37 amino acids at its C-terminus. These considerations led us to hypothesize that mutations in Cx30 might also be responsible for hearing loss. Eight-eight recessive nonsyndromic hearing loss families from both American and Japanese populations were screened for mutations. In addition, 23 dominant hearing loss families and 6 singleton families presumed to be recessive were tested. No significant mutation has been found in the dominant or recessive families.

Amino Acid Sequence↗
PubMed1999

Novel mutation in the KCNQ4 gene in a large kindred with dominant progressive hearing loss.

Analysis of genotyping of a five-generation American family with nonsyndromic dominant progressive hearing loss indicated linkage to the DFNA2 locus on chromosome 1p34. This kindred consists of 170 individuals, of which 51 are affected. Pure tone audiograms, medical records, and blood samples were obtained from 36 family members. Linkage analysis with five microsatellite markers spanning the region around DFNA2 produced a lod score of 6.6 for the marker MYCL1 at straight theta = 0.0. Hearing loss in this family showed a very similar pattern as the first reported American family with the same linkage. High frequency hearing loss was detectable as early as 3 years of age, and progressed to severe to profound loss by the fourth decade. Using intronic primers, we screened the coding region of the KCNQ4 gene. Heteroduplex analysis followed by direct sequencing identified a T-->C transition at position 842, which would produce an L281S amino acid substitution. The observed mutation was shown to segregate completely with affected status in this family. The L281 residue is significantly conserved among the other members of the voltage-gated K(+) channel genes superfamily. Hydrophobicity analysis indicated that L281S substitution would lower formation of the beta structure at the P region of this ion channel. Mutation analysis of KCNQ4 was also performed on 80 unrelated probands from families with recessive or dominant nonsyndromic hearing loss. None of these cases showed a truncated mutation in KCNQ4.

Amino Acid Sequence↗
PubMed1999

Nucleolar ultrastructure in bovine nuclear transfer embryos.

Nuclear transfer experiments in mammals have attempted to reprogram a donor nucleus to a state equivalent to the zygotic one. Reprogramming of the donor nucleus is, among other features, indicated by a synthesis of ribosomal RNA (rRNA). The initiation of rRNA synthesis is simultaneously reflected in nuclear morphology as a transformation of the nucleolus precursor body into a functional rRNA synthesising nucleolus with a characteristic ultrastructure. We examined nucleolar ultrastructure in bovine in vitro produced (control) embryos and in nuclear transfer embryos reconstructed from a MII phase (nonactivated) or S phase (activated) cytoplasts. Control embryos were fixed at the two-, four-, early eight- and late eight-cell stages; nuclear transfer embryos were fixed at 1 and 3 hr post fusion and at the two-, four-, and eight-cell stages. Control embryos possessed a nucleolar precursor body throughout all three cell cycles. In the eight-cell stage embryo, a primary vacuole appeared as an electron lucid area originating in the centre of the nucleolar precursor body. In nuclear transfer embryos reconstructed from nonactivated cytoplasts, the nuclear envelope was fragmented or completely broken down at 1 hr after fusion and, by 3 hr after fusion, it was restored again. At this time, the reticulated fibrillo-granular nucleolus had an almost round shape. The nucleolar precursor body seen in the two-cell stage nuclear transfer embryos consisted of intermingled filamentous components and secondary vacuoles. A nucleolar precursor body typical for the two-cell stage control embryos was never observed. None of the reconstructed embryos of this group reached the eight-cell stage. Nuclear transfer embryos reconstructed from activated cytoplasts, in contrast, exhibited a complete nuclear envelope at all time intervals after fusion. In the two-cell stage nuclear transfer embryo, the originally reticulated nucleolus of the donor blastomere had changed into a typical nucleolar precursor body consisting of a homogeneous fibrillar structure. A primary vacuole appeared in the four-cell stage nuclear transfer embryos, which was one cell cycle earlier than in control embryos. Only nuclear transfer embryos reconstructed from activated cytoplasts underwent complete remodelling of the nucleolus. The reorganisation of the donor nucleolar architecture into a functionally active nucleolus was observed as early as in the four-cell stage nuclear transfer embryo. These ultrastructural observations were correlated with our autoradiographic data on the initiation of RNA synthesis in nuclear transfer embryos.

Animals↗
PubMed1999

Hypoxia limits differentiation and up-regulates expression and activity of prostaglandin H synthase 2 in cultured trophoblast from term human placenta.

OBJECTIVE: We determined the effect of hypoxic conditions on cellular differentiation and prostaglandin H synthase expression in cultured human term trophoblast. STUDY DESIGN: Cytotrophoblasts isolated from term placentas were cultured for 24-72 hours in a standard (20% oxygen) or hypoxic (1% to 2% oxygen) atmosphere. Trophoblast biochemical differentiation was determined by release of human chorionic gonadotropin. Morphologic differentiation was evaluated by epifluorescent and confocal microscopic examination of cultures after dual cytochemical staining for surface membrane desmosomes and intracytoplasmic nuclei. Prostaglandin H synthase 2 expression was determined by Western blot analysis and correlated with enzyme activity by immunoassay of 2 prostaglandin H synthase 2 products, prostaglandin E2 and thromboxane. RESULTS: Human chorionic gonadotropin levels in media and syncytiotrophoblast formation were markedly diminished in trophoblast cultured in hypoxia, compared with trophoblast in control cultures, whereas viability was unchanged. Hypoxia up-regulated expression of trophoblast prostaglandin H synthase 2 but not of prostaglandin H synthase 1 and increased prostaglandin E2 and thromboxane release. CONCLUSIONS: Hypoxia limits differentiation and enhances prostaglandin H synthase 2 expression in cultured villous trophoblast. These responses may account for the cytotrophoblast prominence characteristic of placentas exposed to attenuated oxygen delivery.

Blotting, Western↗
PubMed1999

Interpupillary distance in a normal black population.

This report expands on a study by Pryor [Pryor HB. Objective measurement of interpupillary distance. Pediatrics 1969: 44: 973 977] that related normal values of inner canthal distance (ICD), outer canthal distance (OCD) and interpupillary distance (IPD) for Whites, Asians and Mexican Americans. To date, no similar values have been reported for Blacks. Utilizing a sample (n = 931: 485 males; 446 females) of black people (range, birth 24 years), OCD, ICD, and head circumference (HC) were measured and tabulated. We calculated mean IPD according to Pryor's formulation and report that the general mean OCD and ICD in our sample differed significantly from, and were consistently higher than, Pryor's reported measurements for White males and females at each age level (p < 0.001). However, ICD in our sample was significantly lower at birth in both sexes, appeared to increase at a more rapid rate relative to Whites during the first 3 months of life, and reached and maintained a higher value beyond the age of 3 months, with most age groups showing a significant difference in mean ICD measurements. At each age level, the mean IPD values in Whites and Blacks were significantly higher (p < 0.001). Based upon these findings, we suggest that interpupillary distance of Black children and adults be assessed according to the mean proportions for their race.

Age Factors↗
PubMed1999

Obtaining an accepted Investigational New Drug application to operate an umbilical cord blood bank.

BACKGROUND: The residual blood left in the placenta, previously considered a biologic waste, contains sufficient hematopoietic stem and progenitor cells to consistently engraft at least a small recipient. Over the past several years, more than 500 HLA-matched, related and unrelated, allogeneic cord blood transplants have been performed. Consequently, public and private cord blood banks are being developed to meet future demands. Thus, the definition of a suitable and effective cord blood component needs to be critically defined. In February 1997, the US Food and Drug Administration (FDA) proposed that cord blood banks should operate under an Investigational New Drug (IND) license. STUDY DESIGN AND METHODS: Standard operating procedures were designed using standards from the Foundation for Accreditation of Hematopoietic and Cellular Therapy, the American Association of Blood Banks, and the National Marrow Donor Program and in accordance with current good manufacturing practices. The standard operating procedures were field-tested and submitted to the FDA. RESULTS: Issues of the utmost concern to the FDA dealt with transplant recipient outcome data collection, donor recruitment, sample tracking, the use of unlicensed materials, and the reporting of positive infectious disease results. After three attempts, an IND application was approved. CONCLUSIONS: To obtain approval of an IND application, cord blood banks need a set of standard operating procedures that describe cord blood collection, processing, freezing, and storage. Issues relating to potential cord blood recipient identification, cord blood shipping, and reporting of transplant recipient outcomes are also needed. The IND process provides an opportunity for outside reviewers to make suggestions that may be included in the standard operating procedures.

Blood Banks↗
PubMed1999

Quantitative-trait locus for specific language and reading deficits on chromosome 6p.

Reading disability (RD), or dyslexia, is a complex cognitive disorder manifested by difficulties in learning to read, in otherwise normal individuals. Individuals with RD manifest deficits in several reading and language skills. Previous research has suggested the existence of a quantitative-trait locus (QTL) for RD on the short arm of chromosome 6. In the present study, RD subjects' performance in several measures of word recognition and component skills of orthographic coding, phonological decoding, and phoneme awareness were individually subjected to QTL analysis, with a new sample of 126 sib pairs, by means of a multipoint mapping method and eight informative DNA markers on chromosome 6 (D6S461, D6S276, D6S105, D6S306, D6S258, D6S439, D6S291, and D6S1019). The results indicate significant linkage across a distance of at least 5 cM for deficits in orthographic (LOD = 3.10) and phonological (LOD = 2.42) skills, confirming previous findings.

Adult↗
PubMed1999

Mutations in the KCNQ4 gene are responsible for autosomal dominant deafness in four DFNA2 families.

We have previously found linkage to chromosome 1p34 in five large families with autosomal dominant non-syndromic hearing impairment (DFNA2). In all five families, the connexin31 gene ( GJB3 ), located at 1p34 and responsible for non-syndromic autosomal dominant hearing loss in two small Chinese families, has been excluded as the responsible gene. Recently, a fourth member of the KCNQ branch of the K+channel family, KCNQ4, has been cloned. KCNQ4 was mapped to chromosome 1p34 and a single mutation was found in three patients from a small French family with non-syndromic autosomal dominant hearing loss. In this study, we have analysed the KCNQ4 gene for mutations in our five DFNA2 families. Missense mutations altering conserved amino acids were found in three families and an inactivating deletion was present in a fourth family. No KCNQ4 mutation could be found in a single DFNA2 family of Indonesian origin. These results indicate that at least two and possibly three genes responsible for hearing impairment are located close together on chromosome 1p34 and suggest that KCNQ4 mutations may be a relatively frequent cause of autosomal dominant hearing loss.

Amino Acid Sequence↗