Biomedical subjects

R H Miller

Publications and source records attributed to R H Miller.

At least 19 recordsLinked to original sources

PubMed1992

Lack of protective immunity against reinfection with hepatitis C virus.

Some individuals infected with hepatitis C virus (HCV) experience multiple episodes of acute hepatitis. It is unclear whether these episodes are due to reinfection with HCV or to reactivation of the original virus infection. Markers of viral replication and host immunity were studied in five chimpanzees sequentially inoculated over a period of 3 years with different HCV strains of proven infectivity. Each rechallenge of a convalescent chimpanzee with the same or a different HCV strain resulted in the reappearance of viremia, which was due to infection with the subsequent challenge virus. The evidence indicates that HCV infection does not elicit protective immunity against reinfection with homologous or heterologous strains, which raises concerns for the development of effective vaccines against HCV.

Acute Disease↗

PubMed1992

Sequence analysis of the 5' noncoding region of hepatitis C virus.

We have determined the nucleotide sequence of the 5' noncoding (NC) region of the hepatitis C virus (HCV) genome in 44 isolates from around the world. We have identified several HCV isolates with significantly greater sequence heterogeneity than reported previously within the 5' NC region. The most distantly related isolates were only 90.1% identical. Nucleotide insertions were seen in three isolates. Analysis of the nucleotide sequence from 44 HCV isolates in this study combined with that of 37 isolates reported in the literature reveals that the 5' NC region of HCV consists of highly conserved domains interspersed with variable domains. The consensus sequence was identical to the prototype HCV sequence. Nucleotide variations were found in 45 (16%) of the 282 nucleotide positions analyzed and were primarily located in three domains of significant heterogeneity (positions -239 to -222, -167 to -118, and -100 to -72). Conversely, there were three highly conserved domains consisting of 18, 22, and 63 completely invariant nucleotides (positions -263 to -246, -199 to -178, and -65 to -3, respectively). Two nucleotide domains within the 5' NC region, conserved among all HCV isolates studied to date, shared statistically significant similarity with pestivirus 5' NC sequences, providing further evidence for a close evolutionary relationship between these two groups of viruses. Additional analysis revealed the presence of short open reading frames in all HCV isolates. Our sequence analysis of the 5' NC region of the HCV genome provides additional information about conserved elements within this region and suggests a possible functional role for the region in viral replication or gene expression. These data also have implications for selection of optimal primer sequences for the detection of HCV RNA by the PCR assay.

Base Sequence↗

PubMed1992

In vitro and in vivo characterization of blastemal cells from regenerating newt limbs.

To better characterize the cells involved in newt limb regeneration, blastemal cells from accumulation and differentiation phase blastemas were grown in dissociated cell culture, and their morphology and antigenic phenotype determined using a variety of antibodies directed against intermediate filaments, cell adhesion molecules, and extracellular matrix molecules. In addition to previously described blastemal cell morphologies, many of the cells in these cultures had a round cell body, with an eccentrically placed nucleus and a cytoplasm filled with autofluorescent granules. The majority of accumulation phase blastemal cells labeled with antibodies against GFAP, vimentin, 22/18 as well as with antibodies against NCAM, L-1, laminin, and fibronectin. The majority of differentiation phase blastemal cells had a similar phenotype but lacked expression of vimentin and fibronectin. Comparison of the blastemal phenotype in vitro and in vivo showed similar expression characteristics. However, in differentiation phase blastemas, laminin immunoreactivity was concentrated in specific locations. In addition, the proliferation of cultured blastemal cells is stimulated by the addition of a crude brain extract, consistent with previous studies in vivo and in vitro. Taken together, these observations suggest that dissociated cultures of newt limb blastemal cells provide a suitable model for the analysis of the cell and molecular mechanisms involved in limb regeneration.

Animals↗

PubMed1992

Detection of intrahepatic replication of hepatitis C virus RNA by in situ hybridization and comparison with histopathology.

A nonisotopic in situ hybridization (NISH) assay was used to detect hepatitis C virus (HCV) RNA. A synthetic oligonucleotide complementary to bases 252-301 of the highly conserved 5' noncoding region of the HCV genome was end-labeled by terminal deoxynucleotidyltransferase using digoxigenin-conjugated dUTP. The hybridized oligomer was revealed by an immunohistochemical reaction after incubation with an alkaline phosphatase-conjugated anti-digoxigenin antibody and subsequent amplification with a complex of alkaline phosphatase and anti-alkaline phosphatase antibodies. The intracellular distribution of HCV RNA was monitored in the livers of two chimpanzees experimentally infected with the H strain of HCV and compared with the serum alanine aminotransferase activity, serum HCV RNA, and liver histopathology. Most cells were stained in the cytoplasm as early as 2 days after inoculation, 1 and 2 days, respectively, before the appearance of viral RNA in the serum. The time course of HCV RNA replication was correlated with increases in serum alanine aminotransferase. However, neither one paralleled the appearance of liver cell necrosis nor showed any correlation with the inflammatory response. The NISH signal was not found in liver biopsy specimens taken from these two animals before inoculation with HCV, from chimpanzees with acute hepatitis type A, B, or delta, or from two animals never experimentally infected with any hepatitis agent; moreover, it disappeared when the positive specimens were predigested with RNase and it was not observed after hybridization of positive controls with a labeled oligomer unrelated to HCV RNA. Thus, detection of liver HCV RNA by NISH is a sensitive and specific method for studying HCV replication at the cellular level. Intracellular replication of HCV did not appear to be associated with histopathologic changes in the liver, although the correlation with increases of liver enzyme activity in the serum suggested possible damage to the liver cell membrane.

Alanine Transaminase↗

PubMed1992

Importance of primer selection for the detection of hepatitis C virus RNA with the polymerase chain reaction assay.

We compared four primer sets from conserved regions of the hepatitis C virus (HCV) genome for their ability to detect HCV RNA in a "nested" cDNA polymerase chain reaction assay on sera from 114 anti-HCV antibody-positive individuals from around the world. The different primer sets had equivalent sensitivity, detecting less than 1 chimpanzee ID50 (dose that infects 50%) when tested against reference strain H of HCV. We tested equal amounts of RNA extracted from the serum of each individual with the four primer sets. The set derived from two highly conserved domains within the 5' noncoding (NC) region of the HCV genome, which also share significant similarity with Pestivirus 5' NC sequences, was the most effective at detecting HCV RNA. All samples positive for HCV RNA with any other primer set were also positive with the primer set from the 5' NC region, and the latter was at least 3 times more likely to detect HCV infection than a primer set from within the nonstructural protein 3-like gene region (P less than 0.001). We had no false positive results in greater than 500 negative controls interspersed among the test samples. The 5' NC region primer set detected HCV-specific RNA, verified by high-stringency Southern blot hybridization and DNA sequencing, in 100% of 15 acute and 33 chronic non-A, non-B hepatitis patients from the United States, Europe, and Asia and 10 hepatocellular carcinoma patients from Africa and Asia that tested negative for the hepatitis B virus-encoded surface antigen. In conclusion, use of an appropriate primer set is crucial for detecting HCV RNA in the serum of infected individuals.

Base Sequence↗

PubMed1992

Access to ambulatory care among noninstitutionalized, activity-limited persons 65 and over.

This study examined the impact of income and insurance type on ambulatory care contact use by persons 65 and over who expressed a limitation in their activity. This large group (39% of noninstitutionalized older persons in 1984) had significantly more health problems and ambulatory care contacts than persons not activity-limited. The only previous study on equity in use of physician services among elderly in poorer than average health found relatively little inequality of use due to income and insurance. This study came to the opposite conclusion. Activity-limited persons without Medicare private supplementary insurance, as well as those with supplementary insurance in the bottom and middle of the income distribution, had 15-32% fewer ambulatory care contacts than activity-limited persons with higher income and private supplementary insurance. Particularly striking were the declines in consumption among middle income persons relative to the reference group, indicating that the issue of equity in consumption of health services among older disabled persons affects a much broader group than only the poor and near-poor.

Activities of Daily Living↗

PubMed1992

The precore gene of the woodchuck hepatitis virus genome is not essential for viral replication in the natural host.

A number of naturally occurring hepatitis B virus mutants that cannot synthesize the virus precore protein have been identified. Such mutants have been associated with more severe forms of hepatitis, including fulminant hepatitis. The most common mutation observed is a substitution of G to A in the distal precore gene that converts a codon specifying Trp (TGG) to a termination codon (TAG). Using oligonucleotide-directed mutagenesis, we have produced the same point mutation in the precore gene of an infectious clone of woodchuck hepatitis virus (WHV). Transfection of mutant WHV DNA into the livers of adult woodchucks resulted in replication of the mutant in three of three susceptible animals. Levels of virus replication and transient elevations in liver enzymes in serum were similar to those of adult animals infected with wild-type WHV. Virions, found to possess mutant precore genes by polymerase chain reaction amplification and DNA sequencing, were recovered from the serum of one of the animals and inoculated subcutaneously into neonatal woodchucks. They produced infection in all five animals studied. The level of virus replication in neonatal animals infected with this mutant virus was comparable to that found in neonatal woodchucks infected with wild-type WHV, but none of five woodchucks infected with the precore mutant virus as neonates became chronic virus carriers. It was concluded that the precore gene of the WHV genome is not essential for virus replication in the natural host but may be important for chronic infection.

Animals↗

PubMed1992

Correlation of factor VII activity and antigen with cholesterol and triglycerides in healthy young adults.

Prospective epidemiological studies found that the plasma level of factor VII activity was a risk factor for ischemic heart disease (IHD). Our laboratory previously demonstrated that young adults (mean age, 35 years) at high risk of IHD had significantly higher plasma factor VII activity and antigen levels than did comparable young adults at low risk. To study the relation of factor VII with lipid metabolism in even younger adults (less than 30 years), using standard techniques we measured plasma factor VII activity and antigen, plasma fibrinogen, and fasting serum lipid fractions in healthy male and female subjects who were at low risk of IHD and who were not on medication. Factor VII antigen correlated significantly with total serum cholesterol, fasting serum triglycerides, and high density lipoprotein (HDL) and low density lipoprotein (LDL) cholesterol (p less than 0.01), and factor VII activity correlated with total and HDL cholesterol (p less than 0.05) in the men (n = 132); however, fibrinogen level did not correlate significantly with any lipid level in this group. We found no significant correlation of factor VII activity or antigen with any lipid levels in the women (n = 65). Our data support the hypothesis that control of plasma factor VII level is linked to lipid metabolism in normal physiology in men. Thus, factor VII level may reflect the mechanism by which male gender imparts added risk for IHD, independent of other established risk factors. This study also supports the use of the factor VII antigen assay, a highly reproducible method, in studies of the relation of factor VII to the risk of IHD.

Adult↗

PubMed1992

B-mode ultrasonography of the paranasal sinuses compared with CT findings.

Ultrasound as a diagnostic aid in the evaluation of the sinuses has been a controversial issue. Sensitivities have been reported from 29% to 100% and specificities from 55% to 99%. These wide variations in results from previous studies may have resulted from technical differences. However, the "gold standard" most often used for comparison has been a Water's view radiograph, which has been shown to correlate poorly with intraoperative and CT scan findings. To our knowledge, no one in the English language literature has evaluated sonography of the sinuses with computed tomography as the gold standard. In addition, evaluation of the ethmoid sinus has been neglected by previous studies that have primarily assessed the maxillary sinus. We report the results of a blinded, prospective trial comprised of 41 patients examined by B-mode ultrasound with comparison to computed tomography. Frontal and maxillary sinuses were examined with traditional sonographic techniques and the ethmoid sinuses were scanned with a transglobe technique not previously evaluated in the English language literature. Excluding isolated mucous retention cysts and focal minimal mucosal thickening (defined as < 4 mm), sensitivities for ultrasound of the maxillary, frontal, and ethmoid sinuses were 100% each. Specificities were 98% for the maxillary sinus, 100% for the frontal sinus, and 94% for the ethmoid sinus. Technical aspects and implications of this and previous reports will be discussed.

Adolescent↗

PubMed1992

Correlated response in growth and body measurements accompanying selection for milk yield in Jerseys.

Growth and body measurements from a long-term selection project were analyzed to determine correlated responses to single-trait selection for milk yield. Data were from 1056 daughters (765 selection, 291 control) of 37 bulls (17 selection, 20 control) of 37 bulls (17 selection, 20 control) and included BW and measures of heart girth, chest depth, wither height, and length from withers to pins and from withers to hooks taken at 6 mo, 15 mo, first calving, end of first lactation, and maturity. Other data were birth weight, change in measurements and weights from first calving to end of first lactation, monthly rate of gain from 1 to 13 mo of age, and age reaching breeding weight (250 kg). Principal component scores were calculated from standardized measurements at each age. The first three principal components has meaning (size, length vs. girth, and height vs. girth). All analyses used linear mixed models with fixed effects of genetic group, generation within group, year-season of birth or calving, parity of dam, and birth status (multiple or single birth). Sires were assumed to be random and nested within genetic group. Mean squares for sires was used to test for group differences. Generation did not differ in any analysis and was removed from all models. Selection cows were heavier, larger in some measurements, and had greater overall size at 6 mo of age. Selection cows had greater monthly rate of gain and attained breeding weight at an earlier age. Genetic groups did not differ for any other measurement or weight. Control cows gained more weight and increased more in some measurements between first calving and end of first lactation. Selection for milk yield did not result in an undesirable correlated response in an growth or body measurement.

Animals↗

PubMed1992

Correlated responses in reproduction accompanying selection for milk yield in Jerseys.

Reproductive traits of heifers and primiparous cows from a long-term selection project were analyzed to determine correlated response to single-trait selection for milk yield. Data were from 1056 daughters (765 selection, 291 control) of 37 bulls (17 selection, 20 control). Traits in heifers were ages at first observed estrus and at first breeding, services to conception, interval from first service to conception, and length of first gestation. Traits in primiparous cows were ages at first calving and at first breeding, after calving; services to conception; length of second gestation; and intervals from calving to first observed estrus, to first breeding, and to conception, from first service to conception, and from first to second calving. Analyses for services to conception in heifers and primiparous cows were categorical using models containing genetic group and generation. Analyses of other traits were by linear mixed models using fixed effects of genetic group, generation within group, and year-season of birth. Sires were assumed random and nested within genetic group. The mean square for sires within group was used to test for group differences. No significant differences were found between genetic groups in traits measured in heifers; however, the interval from first service to conception approached significance (control superior). In primiparous cows, differences between genetic groups were significant for the intervals of calving to first breeding and calving to conception and for length of second gestation (control superior). For other traits, reproductive performance of the control was better but not significantly different from that of the selected group. Reproductive performance should be monitored during selection for high milk yield.

Age Factors↗

PubMed1992

Effect of clinical and subclinical mastitis on lipid composition of teat canal keratin.

Two experiments were conducted to investigate lipid composition of teat canal keratin when different conditions of bacterial colonization and quarter inflammation were present. In Experiment 1, 11 multiparous cows with subclinical mastitis (bacteria present but no visible inflammation) in at least one quarter were selected for study. Quarters that were sampled and found negative for bacterial growth were classified as control. In Experiment 2, 10 multiparous cows with clinical mastitis in one or more quarters were selected. Milk samples from inflamed quarters were cultured to identify mastitis pathogens; these quarters were classified as clinical; all other quarters were classified as control. Teat canal keratin was collected from all quarters just before an a.m. milking, samples were weighed, and lipid determinations were conducted by TLC. Keratin from subclinical quarters compared with keratin from control quarters did not differ in either neutral lipid or fatty acid composition. Total lipid was significantly higher in keratin from teats of clinical quarters than in keratin from control quarters (27.8 vs. 21.5 microgram/mg). Neutral lipid composition of keratin was similar between teats from clinical quarters and teats from control quarters. In Experiment 2, quarter foremilk samples were also obtained to determine lipid composition. The FFA in milk from clinical quarters contained fewer short-chain fatty acids, whereas polyunsaturated fatty acids were significantly higher in milk from clinical quarters.

Animals↗

PubMed1992

Influence of involution on intramammary phagocytic defense mechanisms.

Mammary secretions (n = 34 cows) and mammary phagocytes (n = 18 cows) were collected throughout the nonlactating (dry) period to determine changes in intramammary phagocytic defense mechanisms. Mammary secretions were evaluated for their ability to support phagocytosis of Staphylococcus aureus by neutrophils from donor cows and mammary phagocytes for phagocytic and chemiluminescence activity. Ability of secretions to support phagocytosis decreased with advancing length of the dry period. This effect was more pronounced when dry cow secretions constituted 50% of the phagocytic mixture. Phagocytic activity of mammary phagocytes decreased with advancing dry period when autologous secretion was used in the incubation mixture. With homologous secretion, the percentage of phagocytosis increased 5 to 6 d after drying off compared with before drying off and then gradually decreased throughout the remainder of the dry period. Chemiluminescence activity (log10 counts per minute) of mammary phagocytes was lower during the dry period and decreased with advancing dry period. Results indicated diminishing ability of secretions to support phagocytosis and diminished phagocytic and bactericidal mechanisms during the dry period.

Animals↗

PubMed1992

Increased susceptibility to intramammary infection following removal of teat canal keratin.

Influence of teat canal keratin on susceptibility to intramammary infection was investigated in lactating Jersey cows. In each of two replicate trials, keratin was removed from the left teats of 20 cows immediately before milking. Immediately after milking, all teats were exposed to bacterial challenge by immersion in a suspension of Streptococcus agalactiae (5 x 10(7) cfu/ml). Bacterial challenge was repeated after the next four milkings. Foremilk samples were obtained for 8 d after keratin removal to determine infection status. A mammary quarter was classified as infected based solely upon the bacteriological criteria outlined by the National Mastitis Council. The rate of infection in quarters from which keratin was removed was greater than that in control quarters. Infection rates were 26.3% for keratin-removed quarters and 8.3% for control quarters in trial 1 and 13.5 and 0%, respectively, in trial 2. When more stringent criteria (recovery of greater than 100 cfu of S. agalactiae/ml in three or more successive milk samples and a SCC of greater than 10(6)) were used to identify a subset of infections that were clearly intramammary, infection rates were 9.3% for keratin-removed quarters and 1.4% for control quarters. Thus, partial removal of keratin from the teat canal compromised the ability of the teat to prevent passage of bacterial pathogens from the external environment into the mammary gland.

Animals↗

PubMed1992

The American manufacturer's view on the EEC texts and harmonization.

In a world that has become increasingly smaller with the aid of modern air travel, computers and facsimile machines, the European Community's efforts toward harmonization are applauded by the Animal Health Institute, representing the major U.S. manufacturers of veterinary biological products. The new and proposed directives and guidelines are based on the latest technology and help ensure that only high-quality safe and effective veterinary biologicals are registered in the European Community. These new legislations are, however, significantly different from the current United States (USDA) licensing requirements. The USDA requirements are designed to ensure the safety and efficacy of veterinary biologicals licensed in the United States and have been effective in doing so. Among major differences between EEC and USDA requirements are the EEC requirements for GLP safety tests, GMP for manufacturing, with increased measures for the prevention of cross-contamination, duration of immunity data, and raw material specifications in excess of those required in the United States. Very tight deadlines for implementing these requirements have been established. In the spirit of international harmonization and with the goal of reducing or preventing potential trade barriers, the AHI would like to propose the formation of an international working group consisting of FEDESA, AHI, CVMP, USDA and other international industry and government officials. It is suggested that annual meetings be held in conjunction with either FEDESA or AHI meetings. Issues to be dealt with include standardization of test requirements, time lines for implementation and other issues that could reduce or prevent trade barriers without adversely affecting product quality.(ABSTRACT TRUNCATED AT 250 WORDS)

Animal Testing Alternatives↗

PubMed1992

Polysialic acid is required for optimal growth of axons on a neuronal substrate.

Formation of axonal pathways involves a variety of molecules that influence cell-cell interactions. The polysialic acid (PSA) moiety of the neural cell adhesion molecule (NCAM) is present on neuronal surfaces during process outgrowth. Our studies reveal that the removal of PSA causes a decrease in the rate of elongation of retinal cell processes on a substrate of neuronal membranes derived from chick tectum. This effect was partially reversed by antibodies against the L1 adhesion molecule, but not by antibodies against NCAM, N-cadherin, or beta 1-integrins. This predominant effect of PSA on L1 was also observed in short-term, cell-cell adhesion assays, suggesting that PSA promotes optimal outgrowth on neuronal substrates by limiting the consequences of L1-mediated adhesion.

Animals↗

PubMed1992

Cervical lymph node biopsy.

Controversy among head and neck oncologists has centered for many years around the appropriate and inappropriate use of cervical lymph node biopsy. Many authors contend that untimely cervical node biopsy, at best, is unnecessary, and, at worst, may lead to early recurrence and death. More recent data show that adjuvant radiation therapy may ameliorate the adverse effects of lymph node biopsy. At present, although cervical lymph node biopsy is not recommended for the diagnosis of head and neck cancer, it does not appear to shorten survival if appropriate management is administered subsequently.

Biopsy↗

PubMed1992

Astrocyte precursors in neonatal rat spinal cord cultures.

Cultures of newborn rat spinal cord contain multiple types of astrocytes. By using a combination of cultures enriched for glial precursors and clonal analysis, we have identified a particular astrocyte precursor that gives rise to morphologically distinct classes of astrocytes. This astrocyte precursor labels with the monoclonal antibody A2B5, is highly migratory, proliferates in response to serum and platelet-derived growth factor, and differentiates into process-bearing astrocytes, many of which subsequently assume a "pancake"-shaped morphology. A2B5+ astrocyte precursors share antigenic and migratory characteristics with previously described O2A progenitor cells but differ in their response to regulatory factors, including serum and coculture with type 1 astrocytes. More importantly, these astrocyte precursors do not give rise to oligodendrocytes. In their proliferative response to serum and their capacity to differentiate into astrocytes, these glial precursors resemble type 1 astrocyte precursors from optic nerve. However, unlike type 1 astrocyte precursors, these cells are A2B5+, highly migratory, and do not give rise to fibroblast-like astrocytes. Neonatal rat spinal cord cultures contain approximately twice the number of the A2B5+ astrocyte precursors than O2A progenitor cells. By contrast, the majority of A2B5+ cells in postnatal day 7 optic nerve cultures are O2A progenitors. The presence of large numbers of A2B5+ astrocyte precursors in rat spinal cord cultures may reflect the more complex cytoarchitecture of the spinal cord compared to the optic nerve.

Animals↗