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F D Miller

Publications and source records attributed to F D Miller.

At least 73 records · Page 4Linked to original sources
PubMed1994

The T alpha 1 alpha-tubulin promoter specifies gene expression as a function of neuronal growth and regeneration in transgenic mice.

We have previously demonstrated that one member of the alpha-tubulin multigene family, termed T alpha 1 in rats, is regulated as a function of neuronal growth and regeneration. To elucidate the molecular mechanisms responsible for coupling gene expression to morphological differentiation, we have isolated the T alpha 1 gene, have fused 1.1 kb of the 5' flanking region to a nuclear lacZ reporter gene, and have generated transgenic mice. Analysis of these transgenic mice demonstrated that marker gene expression was specific to the CNS and PNS, with expression in vivo at embryonic day 13.5 being similar to expression of the endogenous gene. Moreover, the induction of transgene expression was correlated temporally with neuronal commitment in developing neural crest-derived peripheral neurons and in the developing retina. Immunocytochemical analysis of mixed primary embryonic brain cultures confirmed that transgene expression was specific to neurons, with the majority of neurons, but not astrocytes or oligodendrocytes, expressing beta-galactosidase. Transgene expression in vivo was maintained in developing neurons until early in postnatal life, subsequent to which its expression decreased coincident with neuronal maturation. The transgene was then reinduced in regenerating facial motoneurons following unilateral axotomy of the facial nerve. Thus, 1.1 kb of 5' flanking sequence from the T alpha 1 gene contains the sequence elements responsible for specifying gene expression to embryonic neurons and for subsequently regulating gene expression in both developing and mature neurons as a function of morphological growth.

Animals↗
PubMed1993

Evidence that the loss of homeostatic signals induces regeneration-associated alterations in neuronal gene expression.

Axotomy of motoneurons leads to upregulation of T alpha 1 alpha-tubulin and p75 NGF receptor mRNAs. To distinguish whether these increases are due to interruption of ongoing homeostatic signals or to positive factors derived from non-neuronal cells of the injured nerve, we developed an experimental paradigm that allowed us to selectively block some facets of axonal signaling without initiating a peripheral nerve injury response. More specifically, the marginal mandibular and buccal branches of the facial nerve were locally cooled to 4-8 degrees C (a cold block) for 36-60 hr. This treatment effectively and reversibly blocked fast axonal transport, as monitored by the ability of facial motor neurons to retrogradely transport fluorogold from their terminals to their cell bodies. Light microscopy of semithin epoxy sections demonstrated that the cold block treatment did not lead to macrophage invasion or to morphological alterations in the nerve either proximal or distal to the cold block region. In situ hybridization and image analysis of retrogradely labeled facial motor neurons indicated that such a cold block induced T alpha 1 alpha-tubulin and p75 NGF receptor mRNAs to the same level as did a corresponding nerve transection. In contrast, T26 alpha-tubulin mRNA, which does not increase following axotomy, was not affected by the cold block treatment. These results suggest that neurons sense their status by a constant flow of information carried, circulated, or maintained by fast axonal transport and that the axotomy-induced increases in T alpha 1 and p75 NGF receptor mRNAs in motor neurons are, to a great extent, due to loss of such homeostatic signals.

Animals↗
PubMed1993

Induction of T alpha 1 alpha-tubulin mRNA during neuronal regeneration is a function of the amount of axon lost.

Axotomy of mature peripheral neurons leads to upregulation of T alpha 1 alpha-tubulin mRNA. To distinguish whether this increase is due to interruption of target contact or to loss of the axon itself, we developed a model system that allowed us to axotomize mature sympathetic neurons proximal or distal to their cell bodies, severing contact with target tissue in all cases. Sympathetic neurons within the same superior cervical ganglion that project via the internal and external carotid nerves to the eye and to the ear, respectively, were differentially labeled with the retrograde tracers fast blue and fluorogold, and the labeled neurons were then transected close to or far from their cell bodies. In situ hybridization analysis 5 days postaxotomy (the time of peak T alpha 1 mRNA expression) indicated that when eye and ear neurons were both axotomized close to their cell bodies. T alpha 1 mRNA increased to a similar degree in both populations. In contrast, when ear neurons were again cut close, but eye neurons of the same ganglion were transected far from their cell bodies by eye removal, peak T alpha 1 mRNA levels were two- to threefold lower in the eye neurons. Thus, the increase in T alpha 1 mRNA was much lower when only a small amount of axon was lost, even though contact with target tissue was completely interrupted in all groups. These results therefore suggest that neurons normally monitor the status of their axon via a mechanism that allows for graded responses and that the axotomy-induced increase in T alpha 1 mRNA is, to a great extent, due to loss of the nonterminal axon.

Animals↗
PubMed1993

Gene expression in the developing cerebellum during perinatal hypo- and hyperthyroidism.

The intensity of p75NGFR receptor-like immunoreactivity and the mRNAs encoding p75NGFR, T alpha 1 alpha-tubulin, GAP-43 and the myelin proteins MBP and PLP were measured in the developing cerebellum to study the effects of perinatal thyroid hormone imbalance in rats. Results compared to age-matched controls provide in vivo evidence for differential gene regulation by thyroid hormone in the developing cerebellum. We found that p75NGFR immunoreactivity was strikingly elevated in hypothyroid rats, whereas p75NGFR mRNA content remained only twice as high as that of control levels on postnatal day 15 (P15). When p75NGFR immunoreactivity was still elevated in hypothyroid rats, Purkinje cells exhibited proximal axonal varicosities, axonal twisting and differences in axonal caliber. The mRNAs encoding proteins involved with neurite growth-promoting elements, T alpha 1 alpha-tubulin and GAP-43, were also increased in hypothyroidism, possibly reflecting a neuronal response to a deficiency in, or damage to, cerebellar neurons, or a general delay in their down regulation. Similar increases were not observed for the myelin specific genes. MBP and PLP mRNAs were first detected on P2 of hyperthyroid rats, and they increased with age. Hypo- or hyperthyroidism did not affect the initial onset of MBP and PLP expression, however, hyperthyroidism increased levels of PLP and MBP mRNAs between P2 and P10. By contrast, the most consistent decrease in MBP and PLP mRNAs in rats with thyroid hormone deficiency was observed only on P10. At later times (P15 and P30), the two mRNA levels were similar to controls in all groups. These results are consistent with a role for thyroid hormone in the earlier stages of cerebellar myelination. Hypothryoidism led to specific increases in T alpha 1 alpha-tubulin and GAP-43 mRNAs, and in the immunoreactivity and mRNA levels of p75NGFR receptor--all changes that may play a role in the observed abnormal neuronal outgrowth.

Animals↗
PubMed1993

Mortality and morbidity among blue and white collar workers in the Honolulu Heart Program cohort.

There has been a controversy regarding occupation, as defined by blue and white collar work, and coronary heart disease (CHD). In a large long-term prospective study of the Honolulu Heart Program cohort, there were no significant differences between these two occupational classifications and the incidence rates of fatal and non-fatal CHD. Nor were there significant differences in total mortality, stroke or total cancer although there were very small but significant differences in some CHD risk factors. Unlike the previous studies, this investigation was free of prevalent cases of CHD at baseline and controlled for the major risk factors of CHD. We question the utility of using this occupational classification to investigate the causes of CHD.

Aged↗
PubMed1993

Synthesis of oxytocin in amnion, chorion, and decidua may influence the timing of human parturition.

Despite the widespread clinical use of oxytocin (OT) as a potent and specific stimulant of labor, previous research data have not supported a role for OT in the physiology of normal human parturition. We have demonstrated synthesis of OT mRNA in amnion, chorion, and decidua using Northern blot analysis, ribonuclease protection assays, and in situ hybridization. Probes directed towards both the 3' and 5' ends of the gene have been used. Levels were highest in decidua with considerably less in chorion and amnion and very low levels in placenta. The transcript size in decidua appears to be 60-80 nucleotides smaller than the transcripts in amnion and chorion. OT gene expression in chorio-decidual tissues increased three- to fourfold around the time of labor onset. Estradiol stimulated synthesis of OT mRNA during in vitro incubation. These results support the hypothesis of a paracrine system involving OT and sex steroids within intrauterine tissues wherein significant changes could occur without being reflected in the maternal circulation. Such a paracrine system could rationalize a long-sought role for oxytocin in the physiology of human labor. These data may lead to novel approaches towards prevention or treatment or preterm labor.

Amnion↗
PubMed1993

An epidemiological assessment of Ambrosia maritima on the transmission of schistosomiasis in the Egyptian Nile Delta.

Ambrosia maritima (Damsissa), a proven molluscicide, was investigated in a seven year epidemiological trial in four villages in the northern Egyptian Nile Delta. Schistosoma mansoni prevalence and other measures of infection were initially high in the four villages before the trial began. Two villages were used to test the impact of A. maritima application both on snail populations and on infection in the village population. Two villages were held as controls and not treated with A. maritima. The entire population of all four villages was included in the study. Prevalence and other measures of infection fell dramatically following treatment with praziquantel 40 mg kg-1 body weight. On annual follow ups, the prevalence of infection and geometric mean egg counts began to increase back to original levels in both test and control villages; age adjusted incidence rates were lower in one test village, but higher in the other when compared to the control villages. Snail populations were destroyed in the treated canals and drains located near the test villages. The lack of a clear epidemiologic impact is discussed.

Adolescent↗
PubMed1993

Sugarcane workers: morbidity and mortality.

Sugarcane is, after pineapple, the largest agricultural industry in Hawaii. There have been reports that this industry poses certain health hazards. To investigate this possible hazard in Hawaii, the relationship of employment on a sugarcane plantation to total mortality, the development of definite coronary heart disease (CHD), stroke, cancer, lung cancer and certain risk factors were examined in men of Japanese ancestry participating in the Honolulu Heart Program. After 18 years of follow-up, those men who indicated one or more years working on sugarcane plantations had no significant difference in age-adjusted mortality, nor incidence of CHD, stroke, cancer, or lung cancer. There were no differences in risk factors compared to participants who were never employed on sugarcane plantations, nor were there differences in lung function as measured by FEV1. These findings were unchanged after adjusting for several potential confounding variables. No cases of mesothelioma were observed among those with a history of defined exposure. These findings were not due to a "healthy worker bias" and indicate that employment on a sugarcane plantation in Hawaii is not associated with elevated rates of chronic diseases.

Agricultural Workers' Diseases↗
PubMed1992

Substrate specificity of isopenicillin N synthase.

Highly purified isopenicillin N synthase (IPNS) from two sources (naturally occurring in Penicillium chrysogenum and that expressed in Escherichia coli via a cloned gene derived from Cephalosporium acremonium) have been isolated and utilized in vitro to test synthetic modifications of the natural substrate, (L-alpha-amino-delta-adipyl)-L-cysteinyl-D-valine (ACV). A very sensitive procedure utilizing the ability of beta-lactams to induce the synthesis of beta-lactamase was employed to determine whether an ACV analogue could serve as a substrate for IPNS. A wide variety of amino and carboxyl terminal tripeptide substitutions were examined and found to elicit positive beta-lactamase induction profiles. However, none of these modifications were found to function as efficiently as a substrate as ACV. One of the beta-lactam products which was formed from the reaction of IPNS and the tripeptide analogue was independently synthesized and evaluated for antibacterial activity. Modification of the L-cysteine residue in the second position of ACV resulted in tripeptides that were unable to serve as substrates. Conversion of the D-valine residue in the third position of ACV to an aromatic amino acid or to a highly electronegative residue such as trifluorovaline resulted in elimination of substrate activity and creation of an inhibitor of the enzyme.

Amino Acid Sequence↗
PubMed1992

Concentration-dependent regulation of neuronal gene expression by nerve growth factor.

NGF is a neurotrophic protein that promotes the survival, growth, and differentiation of developing sympathetic neurons. To directly determine the effects of different concentrations of NGF on neuronal gene expression, we examined mRNAs encoding the p75 low-affinity NGF (LNGF) receptor, T alpha 1 alpha-tubulin (T alpha 1), and tyrosine hydroxylase (TH) in pure cultures of rat sympathetic neurons from postnatal day 1 superior cervical ganglia. Studies of the timecourse of gene expression during 2 wk in culture indicated that a 5-d incubation period would be optimal for the concentration-effect studies. Analysis of RNA isolated from neurons cultured in 2-200 ng/ml 2.5S NGF for 5 d revealed that, as the NGF concentration increased, neurons expressed correspondingly increased levels of all three mRNAs. Both LNGF receptor and TH mRNAs increased seven-fold, and T alpha 1 mRNA increased four-fold in neurons cultured in 200 versus 10 ng/ml NGF. In contrast, T26 alpha-tubulin mRNA, which is constitutively expressed, did not alter as a function of NGF concentration. When neurons were initially cultured in 10 ng/ml NGF for 5 d, and then 200 ng/ml NGF was added, LNGF receptor, T alpha 1, and TH mRNAs all increased within 48 h. The timecourse of induction differed: T alpha 1 mRNA was maximal by 5 h, whereas LNGF receptor and TH mRNAs first began to increase at 12 h after the NGF increase. These experiments show that NGF regulates expression of a subset of mRNAs important to neuronal growth and differentiation over a broad concentration range, suggesting that the effects of NGF may be mediated by more than just a single receptor operating at one fixed affinity. These results also suggest a mechanism for coupling neuronal synthesis of axonal proteins to increases in size of the innervated target territory during growth of the organism.

Animals↗
PubMed1992

Spatiotemporal increases in epidermal growth factor receptors following peripheral nerve injury.

Non-neuronal cells of peripheral nerve respond to axonal injury with a series of cellular changes that facilitate neuronal regeneration. To characterize the potential role of the epidermal growth factor (EGF) family of proteins in this response, we monitored the expression of EGF receptor mRNA and protein in the injured rat sciatic nerve. EGF receptor mRNA is synthesized in both primary cultured fibroblasts and Schwann cells, and Schwann cells express EGF receptor-like immunoreactivity. In situ hybridization and immunocytochemistry revealed that EGF receptor mRNA and protein are expressed in Schwann cells and fibroblasts of the sciatic nerve in vivo, and that receptor levels increase following nerve injury. Thirty-six hours postlesion, EGF receptors were expressed in gradients along the nerve both proximal and distal to the lesion, with the highest levels localized adjacent to the transection site. By 72 hr, receptor levels were maintained in a gradient in the proximal segment, but were uniformly increased throughout the portions of the distal segment that were analyzed. These changes were similar to those observed for low-affinity NGF receptor mRNA and protein, with transection causing increased expression in both Schwann cells and fibroblasts. Northern blots confirmed that primary cultured fibroblasts express low-affinity NGF receptor mRNA. To determine whether spatiotemporal gradients were a general characteristic of the nerve injury response, we monitored expression of the mRNA encoding the major myelin protein P0. Levels of P0 mRNA decreased initially in cells immediately adjacent to the transection site and, by 72 hr, were uniformly decreased throughout the distal segment. These data suggest that members of the EGF family of proteins may play a role in the peripheral nerve response to injury, and demonstrate a generalized gradient of cellular responses that commence at the transection site and progress distally in the nerve in the absence of intact axons.

Animals↗
PubMed1991

Generation of the truncated form of the nerve growth factor receptor by rat Schwann cells. Evidence for post-translational processing.

These studies were initiated to determine whether the soluble, truncated form of the nerve growth factor (NGF) receptor arises from post-translational processing of the intact, membrane-bound receptor or from an alternatively spliced mRNA. Pulse-chase analysis of cultured primary rat Schwann cells coupled with immunoprecipitations using antibodies to the intracellular and extracellular domains of the receptor were used to monitor receptor production. Three forms of the NGF receptor (80, 83, and 85 kDa) displaying a precursor product relationship were detected over the 2-h chase period; only the 85-kDa species was detected on the cell surface. Truncated receptors (50 and 52 kDa) were detected in conditioned media 5 h after cell labeling but were never observed intracellularly. Polymerase chain reaction and RNase protection analyses of NGF receptor mRNA targeted toward the coding region for the transmembrane domain detected no splice variants that could generate truncated receptor, and media conditioned by fibroblasts transfected with rat receptor cDNA, in which splicing cannot occur, nonetheless contained the truncated receptor protein. Taken together, these results suggest that the truncated NGF receptor does not arise as a distinct translation product but rather from a post-translational modification of the intact, surface-bound form of the protein.

Amino Acid Sequence↗
PubMed1991

Inhibition of protein kinase C by calphostin C is light-dependent.

Calphostin C, a secondary metabolite of the fungus Cladosporium cladosporioides, inhibits protein kinase C by competing at the binding site for diacylglycerol and phorbol esters. Calphostin C is a polycyclic hydrocarbon with strong absorbance in the visible and ultraviolet ranges. In characterizing the activity of this compound, we unexpectedly found that the inhibition of [3H]phorbol dibutyrate binding was dependent on exposure to light. Ordinary fluorescent light was sufficient for full activation. The inhibition of protein kinase C activity in cell-free systems and intact cells also required light. Light-dependent cytotoxicity was seen at concentrations about 5-fold higher than those inhibiting protein kinase C.

Animals↗
PubMed1991

Detection of brain-derived neurotrophic factor-like activity in fibroblasts and Schwann cells: inhibition by antibodies to NGF.

mRNA coding for brain-derived neurotrophic factor (BDNF) has been detected in cultured L929 fibroblasts, rat dermal fibroblasts, and sciatic nerve Schwann cells, as well as in rat skin. Medium conditioned by cultured fibroblasts and Schwann cells also stimulates neurite growth from retinal explants and promotes the survival in culture of BDNF-responsive sensory neurons; biological activity is abolished by antibodies raised against NGF. These results suggest that molecules with BDNF-like activity may be produced by cells in the peripheral nervous system and that the BDNF-like activity in fibroblasts and Schwann cells is derived from molecules immunologically related to NGF. In support of this concept, antibodies against NGF have been found to reduce the biological activity of recombinant BDNF in culture and to cross-react with BDNF on Western blots.

Animals↗
PubMed1991

Regulation of nerve growth factor receptor gene expression by nerve growth factor in the developing peripheral nervous system.

Nerve growth factor (NGF) is a target-derived neurotrophic protein that promotes the survival and growth of developing sympathetic and sensory neurons. We have examined NGF receptor gene expression in these neurons after NGF administration. Northern blot and in situ hybridization analyses demonstrated that NGF given systemically to neonatal rats increased levels of NGF receptor mRNA in sympathetic neurons within the superior cervical ganglion. This increase was accompanied by a differential regulation of genes associated with neurotransmitter phenotype; tyrosine hydroxylase mRNA was increased, but neuropeptide Y mRNA was not. NGF receptor mRNA levels were also increased in L4-L5 dorsal root ganglia, although this mRNA was not expressed uniformly in sensory neurons of control or NGF-treated animals. Levels of T alpha 1 alpha-tubulin mRNA, a marker of neuronal growth, also increased. In contrast to developing neurons, systemic NGF did not increase NGF receptor mRNA in nonneuronal cells of the sciatic nerve. To determine if NGF regulated NGF receptor gene expression at the transcriptional level, we examined PC12 cells. NGF treatment for 6 h increased NGF receptor mRNA fourfold; this increase was inhibited by cycloheximide. Nuclear run-off transcription assays demonstrated that the increase in steady-state NGF receptor mRNA levels was mediated at the transcriptional level. In contrast, although NGF treatment increased steady-state tyrosine hydroxylase mRNA levels, this effect was not blocked by cycloheximide, and was not due to increased transcription. These data raise the possibility that transcriptional regulation of NGF receptor gene expression by target-derived NGF could be a molecular mechanism for potentiating NGF's effects on neurons during developmental periods of neuronal competition and cell death.

Adrenergic Fibers↗
PubMed1991

Solid waste scavenger community: an investigation in Bangkok, Thailand.

A solid waste scavenger community at On-Nooch dump site in Bangkok was investigated. The purpose was to identify the dimensions of the public health risk to this group of people and their community due to exposure to hazardous conditions from waste materials. A cross-sectional descriptive study utilizing field surveys and measurements was performed. The demographic, socioeconomic, health related and environmental characteristics of this community were examined. Health complaints and injuries were inventoried among scavengers. Prevalence of childhood respiratory illness was high especially in those households where smoking was present. Intestinal helminthic and protozoan infection in children were detected. Six individuals with possible HIV infection and a number of Hepatitis B anti-genemia were found among male respondents. An appreciable proportion of respondents fell below normal when tested for lung function. Air pollution measurements showed acceptable ambient air levels except for particulate matters. Water quality was low for both potable and nonpotable water.

Adolescent↗