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Biomedical subjects

J K Smith

Publications and source records attributed to J K Smith.

At least 19 recordsLinked to original sources
PubMed1998

Sequencing using capillary electrophoresis of short tandem repeat alleles separated and purified by high performance liquid chromatography.

Polymerase chain reaction (PCR) amplified alleles need to be isolated and purified before carrying out additional analysis to confirm sequence, number of repeats and microvariants within a short tandem repeat (STR) locus. Also, PCR amplification of tetranucleotide repeat loci, used in DNA typing assays, often result in heteroduplex formation, adding to the complexity of analysis. Sequencing reactions require single specific target DNA for reliable sequencing analysis. Alkylated poly(styrene-divinylbenzene) columns at elevated temperature and gradient elution conditions increase the efficiency of separation to allow for the purification of PCR products. Using the separation technique of ion-pairing reverse-phase (IPRP) high performance liquid chromatography (HPLC), molecular biologists can separate and purify DNA fragments without alteration to the double-stranded DNA sequencing properties. In this study, the IP-RP chromatography technique has been demonstrated by separation of alleles of the short tandem repeat loci of TH01, vWA31, F13A01 and FES/ FPS. Alleles differing in size range of 12 to 4 base pairs were separated by IPRP/HPLC and individual alleles were peak-captured, then cycle-sequenced. These HPLC fractions required no additional steps prior to cycle sequencing. Capillary electrophoresis (CE) was used to sequence the alleles. Furthermore, CE offers advantages over traditional slab methods via automation and higher applied voltages. Interestingly, unlike traditional gel electrophoresis, samples were introduced into the sieving matrix by electrokinetic injection, which allows for multiple injections from a single sample, a key feature for method development. Applied voltage was 320 V per centimeter using a nonderivatized fused silica capillary with an interior diameter of 50 microm and a total length of 47 centimeters. The total analysis time including capillary filling and pre-electrophoresis was less than 30 min for a 220-bp fragment. A sequencing rate of 530 bp/h was achieved using these conditions. By combining the techniques of HPLC separation and CE sequencing, the results confirmed the sequence and number of nucleotide repeats for each STR loci. An average sequencing efficiency of 97% was achieved. Additionally, this method defined the absence of a 9.3 microvariant for a TH01 heterozygous individual previously typed as a 9, 9.3/10 using slab gel electrophoresis. The techniques described can be applied to other DNA purification and isolation problems.

Alleles↗
PubMed1998

The influence of postweaning housing conditions on drug-induced conditioned taste aversion.

Postweaning social isolation can influence the sensitivity of rats to several effects of drugs of abuse. The present study investigated the influence of postweaning housing conditions on the sensitivity of rats to the aversive effects of a number of psychoactive agents using a conditioned taste aversion (CTA) test procedure. Development of a CTA was assessed by pairing administration of the drug with the consumption of a 0.05% (weight/volume) saccharin solution in water-deprived (18 h) rats in a 20 min drinking period. Saccharin consumption was then measured in 20 min test sessions over the next 4 consecutive days. Consumption of saccharin solution was significantly reduced in both isolated and enriched rats following administration of d-amphetamine (2 mg/kg), cocaine (30 mg/kg), morphine (10 mg/kg), nicotine (1.0 mg/kg), caffeine (20 mg/kg), alcohol (1.5 g/kg), and LiCl (0.15 M, 4 ml/kg). There was no significant effect of housing conditions on the CTA induced by cocaine, nicotine, alcohol, or LiCl; however, isolation-reared rats were found to be less sensitive to the aversive effects of d-amphetamine, morphine, and caffeine in this paradigm. These results suggest that rearing rats in social isolation induces an attenuation in sensitivity to the aversive effects of some psychoactive agents.

Animals↗
PubMed1998

Rapid PCR for identity testing using a battery-powered miniature thermal cycler.

A microfabricated, battery-powered thermal cycler was implemented in PCR-based DNA typing for human identification. HLA DQ alpha and an STR triplex were PCR amplified using a device known as the Miniature Analytical Thermal Cycling Instrument (MATCI). The extremely efficient heating properties of the MATCI enabled thermal cycling to be completed in as little as 21 min. In addition, the feasibility of using the real-time fluorescent detection system of the MATCI was demonstrated. The successful application of this portable, prototype device to forensic identity testing is a significant milestone towards the eventual development of a completely integrated DNA testing instrument that would also incorporate sample preparation and allele detection.

DNA↗
PubMed1997

Post-weaning housing conditions influence the behavioural effects of cocaine and d-amphetamine.

Post-weaning social isolation can induce profound and long lasting effects on an animal's behaviour. The present study investigated the influence of post-weaning housing conditions on the sensitivity of rats to the behavioural effects of d-amphetamine and cocaine. The locomotor stimulant effects of both drugs were compared following acute and chronic administration. The influence of post-weaning housing conditions on the effects of d-amphetamine and cocaine on responding for food and for a conditioned reinforcer were also examined. Isolated rats showed enhanced locomotor activity on exposure to a novel environment. This difference was further exaggerated following administration of d-amphetamine (0.5 mg/kg) and cocaine (5 mg/kg). Isolated, but not enriched, rats exhibited sensitisation to the locomotor activating effects of repeated administration of a dose of 0.5 mg/kg d-amphetamine, whilst both groups sensitised equally to a dose of 1.0 mg/kg d-amphetamine. Rearing conditions did not affect sensitisation to cocaine (5, 10 mg/kg). Isolated rats exhibited a higher rate of responding for a conditioned stimulus and for food on a progressive ratio schedule of reinforcement, both of which were enhanced to a greater extent in isolates following administration of cocaine (5 mg/kg) and d-amphetamine (0.5 mg/kg). These results suggest that isolation rearing induces an enhancement in sensitivity to both the locomotor stimulant and reinforcing properties of amphetamine and cocaine.

Animals↗
PubMed1997

Bidirectional effects of dopamine D2 receptor antagonists on responding for a conditioned reinforcer.

In general, the administration of dopamine (DA) antagonists has been shown to result in the attenuation of reward processes. Recently, however, it has been suggested that low doses of DA antagonists can enhance the incentive value of a primary reinforcer. The present study examined the effect of DA receptor antagonists on responding for a conditioned stimulus (CS) and compared their effects to that produced by d-amphetamine. For 12 days, food-deprived rats were trained to associate a CS with a food reward. In the test phase, the CS was presented following a response on one of two levers (CR), whereas responding on the other lever (NCR) had no consequence. Low doses of d-amphetamine (0.5 mg/kg), sulpiride (4 mg/kg), pimozide (0.025 mg/kg), and raclopride (0.05 mg/kg) selectively enhanced responding on CR. A low dose of haloperidol (0.01 mg/kg) led to a nonspecific increase in lever responding. Treatment with larger doses of these compounds as well as with the D1 antagonist SCH23390 attenuated responding on CR. Both CR and NCR responding were reduced following administration of higher doses of d-amphetamine. Results indicate that responding for a conditioned reinforcer is potentiated following administration of low doses of D2 receptor antagonists, suggesting that D2 receptor blockade can facilitate incentive motivation.

Amphetamine↗
PubMed1997

Computed model of gadolinium enhanced MRI of breast disease.

OBJECTIVE: Investigation has shown that the most useful MRI finding for the detection of breast cancer is enhancement following in travenous contrast. However, many widely different imaging protocols have been used. The purpose of this study is to explicate factors that affect the signal intensity of breast lesions after intravenous gadolinium. METHODS AND MATERIALS: A computer model was developed using equations based on published data. The effect of gadolinium on breast tissues was calculated using the model with appropriate values for baseline tissue relaxation times, relaxivity of gadolinium at the given field strength and concentration of gadolinium based on published data, for the TR, TE, flip angle and field strength of several published sequences used for enhanced breast MRI. RESULTS: The computer model allows comparison of the performance of different sequences, which can be displayed graphically. These vary in their performance, largely dependent on T1 weighting. Enhancement is also affected by the baseline of the T1 of the lesion and sensitivity of the sequence to gadolinium. Malignant lesions demonstrate greater observed enhancement than predicted when assuming symmetric distribution of contrast, indicating there is greater accumulation of gadolinium, accounting for the differential enhancement between benign and malignant lesions. CONCLUSIONS: MRI sequences vary greatly in their demonstration of enhancement after intravenous gadolinium contrast. Numerical diagnostic criteria such as % signal intensity change must be interpreted with care when using a different sequence than that on which the criterion was developed. There is preferentially greater accumulation of contrast in malignant lesions, whether due to angiogenesis or altered permeability.

Algorithms↗
PubMed1997

Clinical manifestations of IgE hypogammaglobulinemia.

BACKGROUND: Although IgE has been shown to play a role in the expulsion of intestinal parasites in experimental animals, its overall contribution to host defense in humans remains a subject of controversy. In order to clarify the potential role of IgE in host defense, we have studied the clinical characteristics of patient with serum IgE levels of < 2.5 IU/mL, using patients with normal or elevated IgE levels as controls. OBJECTIVE: To determine the clinical characteristics of IgE deficiency. METHODS: Serum IgE levels were measured in 420 adult patients seen in our Allergy-Immunology Clinic over a period extending from January, 1990 to March, 1996. All subjects were examined by one of the authors (JKS or GHK) using a standardized history and physical examination form. Patients with IgE levels of < 2.5 IU/mL also had measurements of serum IgG, IgG subclasses, IgA and IgM. All IgE-deficient patients and 73% of those with normal to elevated IgE levels underwent RAST and/or skin testing for Type I hypersensitivity, and, where clinically indicated, had serum drawn for autoimmune serologic profiles. Infectious complications were documented by culture. The American Rheumatology Association criteria were used to establish a diagnosis of autoimmune disease. RESULTS: Forty-four patients were found to have IgE levels of < 2.5 IU/mL; 57% of these had depressed serum levels of other immunoglobulins, and 43% had isolated IgE deficiencies. Respiratory symptoms were equally common in IgE-deficient patients and in patients with normal to elevated IgE levels. IgE-deficient patients, however, were more likely to complain of arthralgias (P < .0001), chronic fatigue (P < .0001), and symptoms suggestive of airway infection (P = .0119). Compared with controls, patients with IgE deficiency had a higher prevalence of autoimmune disease (46% versus 15%) (P < .0001) and nonallergic reactive airway disease (73% versus 20%) (P < .0001). There was no difference in the prevalence of these disease in patients with selective IgE deficiency as compared with those with IgE deficiency complicated by deficits in other immunoglobulin classes. IgE-deficient patients with multiple immunoglobulin deficiencies, however, were more likely to have serious infection involving both the upper and lower respiratory tract than those with isolated IgE deficiency. CONCLUSIONS: IgE-deficient patients have an increased prevalence of multiple immunoglobulin deficits, autoimmune disease, and nonallergic reactive airway disease when compared with a clinic population of patients with normal to elevated IgE levels.

Adult↗
PubMed1997

Multifunctional cytokine expression by human mast cells: regulation by T cell membrane contact and glucocorticoids.

Human mast cells readily release a variety of mediators, including cytokines, in response to IgE receptor crosslinking, but the mechanisms governing the expression of cytokines are still unclear. Using a human mast cell line, HMC-1, we show expression of cytokine transcripts as early as 2 h after activation with ionomycin and phorbol myristate acetate (PMA). Resting HMC-1 cells expressed transcripts for interleukin-1 receptor antagonist (IL-1RA), IL-2, IL-4, IL-5, GM-CSF, and weakly for IL-8, and stimulation with ionomycin and PMA induced additional transcripts for IL-6 and IL-13 and upregulated expression of IL-8 transcripts. HMC-1 cells secreted IL-4, IL-8, and GM-CSF protein after activation and dexamethasone significantly inhibited the production of these cytokines. Of significance is the finding that the addition of membranes purified from activated T cells to mast cell cultures induced transcripts selectively for IL-8 and none for other proinflammatory cytokines. Flow cytometry revealed that resting HMC-1 cells express CD40, a molecule involved in contact-dependent signaling of monocytes and B cells by T cells. However, activation of HMC-1 by anti-CD40 antibody did not induce IL-8 gene expression or protein production. This study demonstrates that human mast cells are capable of expressing multiple cytokines that can be inhibited by glucocorticoids. It also raises the possibility that T cells may activate mast cell cytokine synthesis by novel contact-dependent mechanisms. This phenomenon of T cell regulation of mast cell function requires further study.

Anti-Inflammatory Agents↗
PubMed1997

A new flap for nasal tip reconstruction.

Aesthetic reconstruction of tumor defects of the nasal tip and supratip areas remains a challenge. Because of our dissatisfaction with the current reconstructive options, we have developed a modification of the nasalis flap based on the angular artery for the management of nasal tip defects. This is an axial flap-pattern that rotates toward the midline and nasal tip, leaving donor scars in the nasojugal and alar creases. A second rotation flap from the alar groove fills the donor defect in the nasojugal region to minimize scarring. We have utilized this flap 19 times in 18 consecutive patients with nasal tip and supratip tumor defects with follow-up ranging from 10 to 72 months. Donor scars are well concealed and nasal contour is minimally altered. Color and texture matches are excellent, and the pincushion deformity has been eliminated. On the basis of these results, we believe the modified nasalis flap is ideally suited for nasal tip reconstruction.

Aged↗
PubMed1997

Studies on the inhibitory effects of analogues of dapsone on neutrophil function in-vitro.

We have compared twelve sulphone analogues of dapsone in terms of inhibition both of zymosan-mediated human neutrophil respiratory burst and inhibition of interleukin-1-stimulated neutrophil adhesion to transformed human umbilical vein endothelial cells. Overall, there was a good correlation between the respective rank orders of compound potency in the two test systems. The most effective compounds in terms of respiratory burst and adherence inhibition were the 2-nitro-4-amino-, 2-hydroxy-4-aminopropyl-, and 2-methoxy-4-aminoethyl- derivatives. In general, potency was inversely associated with lipophilicity; compounds with bulky side-chains, e.g. the 2-methyl-4-aminopentyl, 2-methyl-4-aminohexyl and the 2-hydroxymethyl-4-aminoethyl derivatives, were less potent. A 2-hydroxy-4-amino- derivative was the exception, however, with low lipophilicity and relatively low potency. All of the compounds tested showed comparable or greater inhibition in both the neutrophil-mediated assays compared with dapsone. Some of the compounds might, because of their good tissue penetration and lower toxicity than dapsone, have the potential to undergo further development.

Adult↗
PubMed1997

Pancreatic fluid collections prior to intervention: evaluation with MR imaging compared with CT and US.

PURPOSE: To evaluate the ability of magnetic resonance (MR) imaging to depict solid debris within pancreatic collections prior to intervention and to help assess drainability, as well as to compare MR findings with those obtained at computed tomography (CT) and ultrasound (US). MATERIALS AND METHODS: Nineteen collections in 18 patients were evaluated with MR imaging, CT, and US prior to drainage. Prospective, blinded interpretations of imaging studies by three independent readers (each interpreted all the images obtained with only one modality) evaluated collection characteristics (debris, consistency, septation, wall thickness, and irregularity) and predicted drainability. Findings were compared with clinical diagnosis and clinical outcome of drainage. RESULTS: MR imaging and CT depicted all collections; US failed to depict two collections. In nine patients with subacute necrotic collections, solid debris was seen in eight (89%) at MR imaging, in two (22%) at CT, and in eight (89%) at US. In seven patients with pseudocysts, debris was seen in two (28%) at MR imaging and in none at CT, as well as in six (100%) of six at US. A collection was defined as "not drainable" on the basis of the depiction of solid necrotic debris more than 1 cm in diameter. With this definition, statistically significant differences between sensitivity and specificity values, respectively, were found for the prediction of actual drainability: MR imaging, 100% and 100%; CT, 25% and 100%; US, 88% and 54%. CONCLUSION: Predrainage MR imaging should be performed in patients with subacute pancreatic collections to avoid infectious complications from unrecognized necrotic debris that cannot be removed with use of standard pseudocyst drainage techniques.

Abscess↗
PubMed1997

Carotid artery stents: early and intermediate follow-up with Doppler US.

PURPOSE: To determine whether ultrasound (US) is a sensitive follow-up method after placement of a carotid artery stent for the detection of significant stenosis, occlusion, and other complications at early and intermediate follow-up. MATERIALS AND METHODS: Doppler US examinations were performed after stent placement in 170 carotid arteries in 119 patients with angiographic correlation. Prospective diagnostic US criteria for stenosis were peak-systolic velocity greater than 1.25 m/sec, internal carotid artery (ICA) to common carotid artery (CCA) peak-systolic velocity ratio of greater than or equal to 3:1, and intrastent doubling of peak-systolic velocity. Retrospective criteria for stenosis were also applied: peak-systolic velocity greater than 1.7 m/sec, ICA end-diastolic velocity greater than 0.4 m/sec, ICA/CCA peak-systolic velocity ratio greater than 2.0, and ICA/CCA end-diastolic velocity ratio greater than 2.4. RESULTS: Eighty-seven immediate and 83 intermediate (average, 7.3 months) follow-up US examinations were performed. Two stent occlusions were detected. One or more prospective US criteria were abnormal in 26 arteries with a stent. One or more retrospective criteria were positive in 47 arteries. Angiography showed corresponding findings, with only one significant stenosis (63%) in the ICA stents. Moderate collapse of a CCA stent was depicted at US. CONCLUSION: Only one significant recurrent stenosis was detected, and no significant stenoses were missed at US. US successfully depicted carotid artery stent occlusion and a moderate stent collapse. Sensitivity in the detection of intrastent stenosis is promising. Further study to refine US criteria in a study with longer term follow-up is needed owing to the lack of significant recurrent stenosis in the intermediate follow-up group.

Aged↗
PubMed1996

Persistent urticarial eruption in an asthmatic patient.

This case demonstrates the importance of recognizing urticarial vasculitis in patients with chronic urticarial eruptions. The salient points in history that point towards the diagnosis of urticarial vasculitis include the presence of painful urticarial lesions that last longer than 24 hours and that heal leaving residual pigmentation. In some cases the urticaria may evolve into palpable purpura. An associated systemic illness that may resemble systemic lupus erythematosus should also suggest the diagnosis, which is established by skin biopsy. Histopathology reveals a leukocytoclastic vasculitis involving postcapillary venules. When associated with systemic vasculitis, the urticaria is likely to be of the hypocomplementemic variety, with immunoglobulin and complement deposition on biopsies, and with serum complement studies demonstrating classical pathway activation, low C1q levels, and anti-C1q precipitins. A variety of agents have been used in the management of urticarial vasculitis, including aspirin, nonsteroidal anti-inflammatory agents, corticosteroids, colchicine, dapsone, hydroxychloroquine, and cytotoxic agents such as cyclophosphamide and azathioprine.

Adult↗
PubMed1996

Lymphoblastoid interferon-alpha inhibits T cell proliferation and expression of eosinophil-activating cytokines.

T cell-derived cytokines, such as interleukin-5 (IL-5) and granulocyte-macrophage colony-stimulating factor (GM-CSF) activate eosinophils, whereas other cytokines, such as tumor necrosis factor (TNF)-alpha and IL-13, determine eosinophil recruitment. Interferon-alpha (IFN-alpha), a leukocyte-derived cytokine, has been shown to have beneficial effects in eosinophil-mediated disorders, such as the hypereosinophilic syndrome and a murine model of allergic asthma, where it inhibited eosinophil recruitment. We tested the hypothesis that IFN-alpha acted in eosinophil-mediated disorders by modulating T cell cytokine expression. Peripheral blood mononuclear cells (PBMC) or human ragweed-specific TH1 (2B8) and TH2 (2D2) T cell clones were cultured in the presence of 5 micrograms/ml of phytohemagglutinin (PHA) or 25 micrograms/ml of antigen Amb a 1 (short ragweed allergen), respectively, and lymphoblastoid IFN-alpha (varying from 0 to 10,000 U/ml). We assessed T cell proliferation by [3H]thymidine incorporation and production of IL-5 and GM-CSF by ELISA. Expression of cytokine transcripts was analyzed by the reverse transcription-polymerase chain reaction technique (RT-PCR). IFN-alpha induced a dose-dependent suppression of T cell proliferation of both PBMC (p < 0.001) and the T cell clones (p < 0.001). IFN-alpha inhibited gene expression of IL-5, GM-CSF, TNF-alpha, and IL-13 in PBMC. Furthermore, IFN-alpha significantly inhibited mitogen-induced and antigen-induced production of IL-5 and GM-CSF. IFN-alpha may benefit eosinophil-mediated disorders by inhibiting T cell function and production of cytokines active on human eosinophils.

Adult↗
PubMed1996

Postmortem MR imaging of lobar cerebral infarction with pathologic and in vivo correlation.

Postmortem magnetic resonance (MR) imaging features of different types of lobar cerebral infarction are correlated with the findings in gross and histologic specimens. The postmortem findings are also correlated with in vivo findings in similar cases selected from teaching files. In acute infarction, white matter vasogenic edema leads to high signal intensity on T2-weighted images and blurring of the gray-white matter junction. Petechial hemorrhage in the cortex results in inhomogeneous signal intensity on T2-weighted images. In laminar necrosis, the hyperintense cortex on T1-weighted images is due not to hemorrhage but possibly to necrosis and the presence of lipid-laden macrophages. In subacute infarction, cortical edema and necrosis may cause the gyral pattern of enhancement. Meningeal inflammation and early fibrosis are probably responsible for meningeal enhancement. In chronic infarction, gliosis and cystic malacia are responsible for the increased signal intensity of white matter on T2-weighted images. Knowledge of the pathologic features of cerebral infarction helps in understanding the MR imaging findings.

Acute Disease↗
PubMed1996

Resolution of the neutropenia of Felty's syndrome by longterm administration of recombinant granulocyte colony stimulating factor.

Felty's syndrome is characterized by neutropenia, splenomegaly, and recurrent infection in patients with rheumatoid arthritis. We used recombinant granulocyte colony stimulating factor (rGCSF) in a patient with Felty's syndrome and recurrent sepsis. rGCSF induced a statistically significant increase in the patient's absolute neutrophil and total white blood cell counts. During 14 months of followup taking rGCSF, disseminated varicella zoster was the only infectious complication. Except mild thrombocytopenia and a transient flare of arthritis, no serious adverse effects occurred. rGCSF may be a safe and effective therapy for Felty's syndrome in selected patients.

Aged↗
PubMed1996

Effect of interferon alpha on HLA-DR expression by human buccal epithelial cells.

/We have studied the effect of interferon alpha (IFN-alpha) on MHC class II expression by human buccal epithelial cells (BEC), and mRNA expression by BEC and mucosal-associated mononuclear cells (MAMC). In 6 experiments, freshly collected BEC were suspended at a concentration of 1.0 x 10(5)/ml in RPMI 1640 and incubated in the presence of 0-10,000 IU/ml of human lymphoblastoid IFN-alpha (HuIFN-alpha). Zero and six hour samples were analyzed by single color flow cytometry using FITC-labeled murine IgG1 monoclonal antibody to HLA-DR. Preparations were also analyzed for expression of cytokine transcripts (IL-2, IL-4, IL-5, IL-6, IL-8, IFN-gamma, GM-CSF) by reverse transcriptase polymerase chain reaction (RT-PCR). Increasing concentrations of IFN-alpha resulted in proportionate increases in the percentage of HLA-DR + BEC (r = 0.7897, p = 0.0627) and in the percentage of HLA-DR+ staining at higher intensities (10(1) to 10(2) log fluorescence intensity) (LFI) (r = 0.4010, p = 0.0424). The percentage of HLA-DR + BEC rose from a mean of 1.5% with no IFN-alpha to 7% with 10,000 IU/ml IFN-alpha (p < 0.05). The percentage of HLA-DR + BEC staining at 10(1) to 10(2) LFI rose from a mean of 8.3% with no added IFN-alpha to 19.2% with 10,000 IU/ml IFN-alpha (p < 0.05). Unstimulated BEC constitutively expressed IL-8 and GM-CSF. IFN-alpha stimulated preparations also expressed IFN-gamma, possibly due to the presence of MAMC, which comprised 2-9% of the total cell population. These data indicated that HuIFN-alpha upregulates MHC class II expression by human BEC, possibly by enhancing IFN-gamma production by MAMC present in the culture preparations.

Cells, Cultured↗