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Biomedical subjects

A M Jensen

Publications and source records attributed to A M Jensen.

At least 19 recordsLinked to original sources

Continuous observation of multipotential retinal progenitor cells in clonal density culture.

All neural cell types in the vertebrate retina, except astrocytes, have been shown to develop from multipotential progenitor cells. It is not known, however, to what extent the progenitor cells are heterogeneous in their developmental potential or to what extent cell-cell interactions versus cell-autonomous factors influence the types of cells they become. To address these issues we developed a clonal-density cell culture system where mouse retinal progenitor cells can survive, divide, and differentiate. We followed the development of clones both by continuous time-lapse video microscopy and by daily microscopic observation. We show that even when cultured at clonal density in a homogeneous general environment, where they cannot contact cells outside their own clone, the retinal progenitor cells vary in proliferative capacity, cell cycle time, and in the cell types that they generate. In addition, we show that under these conditions single progenitor cells can generate both neurons and glia, in which case the neurons almost always develop before glial cells, as is the case in vivo.

Animals

Regulation of lactase-phlorizin hydrolase gene expression by the caudal-related homoeodomain protein Cdx-2.

Lactase-phlorizin hydrolase is exclusively expressed in the small intestine and is often used as a marker for the differentiation of enterocytes. The cis-element CE-LPH1 found in the lactase-phlorizin hydrolase promoter has previously been shown to bind an intestinal-specific nuclear factor. By electrophoretic mobility-shift assay it was shown that the factor Cdx-2 (a homoeodomain-protein related to caudal) binds to a TTTAC sequence in the CE-LPH1. Furthermore it was demonstrated that Cdx-2 is able to activate reporter gene transcription by binding to CE-LPH1. A mutation in CE-LPH1, which does not affect Cdx-2 binding, results in a higher transcriptional activity, indicating that the CE-LPH1 site contains other binding site(s) in addition to the Cdx-2-binding site.

Base Sequence

Expression of Sonic hedgehog and its putative role as a precursor cell mitogen in the developing mouse retina.

We show that Sonic hedgehog and patched are expressed in adjacent domains in the developing mouse retina. Treatment of cultures of perinatal mouse retinal cells with the amino-terminal fragment of Sonic hedgehog protein results in an increase in the proportion of cells that incorporate bromodeoxuridine, in total cell numbers, and in rod photoreceptors, amacrine cells and Muller glial cells, suggesting that Sonic hedgehog promotes the proliferation of retinal precursor cells. These findings suggest that hedgehog and patched are part of a conserved signalling pathway in retinal development in mammals and insects.

Aging

[Prognostic value of interleukin-8 in AIDS-related Pneumocystis carinii pneumonia].

UNLABELLED: We evaluated the significance of interleukin-8 (IL-8) in Pneumocystis carinii pneumonia (PCP). Bronchoalveolar lavage (BAL) fluid and serum was prospectively collected in 76 consecutive HIV-infected patients with a primary episode of PCP, as well as in ten healthy control subjects. Patients were found to have elevated levels of IL-8 in BAL fluid compared to control subjects (p < 0.01). Nine patients died during the course of PCP. Non-survivors had significantly higher IL-8 levels in BAL fluid than survivors (p < 0.05). Furthermore patients with levels of IL-8 in BAL greater then 90 pg/ml (i.e. greater than control subjects) had significantly worse vital prognosis (log rank test, p < 0.05). Thirteen patients required mechanical ventilation (MV), and these patients had significantly elevated levels of IL-8 compared with patients not requiring MV (p < 0.05). IN CONCLUSION: i) IL-8 in BAL fluid correlates to the clinical severity of the pneumonia, and ii) is a predictor of mortality and severe respiratory compromise.

AIDS-Related Opportunistic Infections

[Prevalence of Chlamydia trachomatis among conscripts. A comparative study of urine samples and urethral swabs].

The purpose of the investigation was 1) to determine the prevalence of C. trachomatis among young men and women enrolled in military service, 2) to compare enzyme-immunoassay (EIA) of a urethral swab with a sample of first voided morning urine, 3) to determine frequency of earlier venereal diseases (VD) and actual symptoms of urethritis and cervicitis. EIA positive specimens were confirmed by immunofluorescent microscopy. The investigation comprised 831 men and 80 women (17-26 years). The prevalence was 5.7% (95% confidence limits 4.2-7.4%) and 15% (8-25%) for men and women, respectively (p < 0.005, chi 2). The agreement between the results of the urethral swab and the urine sample was low. Sensitivity and positive predictive value for urethral swab was better than for urine. The handling of urine specimens was more laborious and confirmation more difficult. The percentage of actual symptoms among Chlamydia positive men was 6. The frequency of earlier VD was 7.5% and 10% among men and women, respectively. Treatment with tetracycline was effective, which makes control of eradication unnecessary. We conclude that genital chlamydial infection among young asymptomatic people is common and screening would be desirable. We still recommend urethral swabbing as the routine method until simpler and/or more reliable assays for urine specimens are developed.

Adolescent

Low bone mass and fast rate of bone loss at menopause: equal risk factors for future fracture: a 15-year follow-up study.

The aim of the study was to examine the role of peak bone mass and rate of postmenopausal bone loss for the subsequent risk of osteoporotic fracture. 182 women within 3 years of menopause were followed longitudinally for 15 years. Over the first 2 years, forearm bone mass (single photon absorptiometry) was measured nine times, the rate of bone loss was calculated, and the women were stratified into a group of "fast bone losers" (n = 49) and a group of "normal bone losers" (n = 133). Later, bone mass was also measured in the lumbar spine and hip with dual energy X-ray absorptiometry. At 15 years, the fast losers had significantly lower body weight (4.4 kG; p < 0.05) than the normal losers. Furthermore, the fast losers had significantly increased values of bone turnover (osteocalcin and C-terminal type I collagen breakdown products). In both the forearm, spine, and hip, the fast bone losers had at all sites significantly less bone mass than the normal bone losers (p < 0.001). 23 women had experienced a peripheral (Colles') fracture and 25 a spinal fracture. The fracture groups had generally significantly (p < 0.05) less bone mass than the group without fracture, both in the forearm, spine, and hip and they also had the highest rate of bone loss after menopause (p < 0.05). Baseline bone mass and rate of loss predisposed to the same extent to fractures with ODD's ratios of about 2. If both low bone mass and rate of loss were present, the ODD's ratio increased to about 3. We conclude that fast rate of bone loss and low bone mass are equally important for the risk of fracture. The identification of women at risk of osteoporosis should therefore consider both a measurement of bone mass status, and a determination of the postmenopausal rate of loss.

Bone Density

Does cytomegalovirus predict a poor prognosis in Pneumocystis carinii pneumonia treated with corticosteroids? A note for caution.

OBJECTIVE: To examine the importance of cytomegalovirus (CMV) in bronchoalveolar lavage (BAL) fluid of patients with HIV-associated Pneumocystis carinii pneumonia (PCP) treated with adjunctive corticosteroids (CS). DESIGN: Analysis of clinical data during a 5-year period. SETTING: Department of infectious diseases where clinical and paraclinical data on patients suspected of having PCP have been sampled prospectively. PATIENTS: 148 consecutive patients with a first episode of PCP in a 5-year period. MAIN OUTCOME MEASURE: Vital status 3 months after diagnosis of PCP. RESULTS: Patients with PCP treated with adjunctive CS who had CMV cultured from BAL fluid had a two times higher mortality within 3 months from bronchoscopy than others (p = 0.08). This difference could not be explained by differences in CD4 count, PO2 or PCO2 at time of bronchoscopy. CONCLUSION: With the accepted usage of adjunctive CS in severe PCP, the role of CMV as a pulmonary copathogen may have changed. Active CMV infection may be an important cause of failing treatment of severe PCP in those treated with adjunctive CS.

AIDS-Related Opportunistic Infections

Overexpression of mitogen-activated protein kinase kinase (MAPKK) and its mutants in NIH 3T3 cells. Evidence that MAPKK involvement in cellular proliferation is regulated by phosphorylation of serine residues in its kinase subdomains VII and VIII.

Mitogen-activated protein kinase kinase (MAPKK) is a dual specificity protein kinase that exhibits a high degree of specificity toward its downstream target, mitogen-activated protein kinase (MAPK). In this study, we used stable overexpression of MAPKK and its mutants in NIH 3T3 cells to study effects on downstream components of the MAPK signaling cascade and to correlate them to physiological responses. We have mutated the potential regulatory serine residue 222 to alanine (S222A) or to glutamate (S222E) and serines 212 and 218 together to alanine residues (S212A,S218A). Lysine 97 was mutated to alanine (K97A) to provide an inactive enzyme. Overexpression of the wild type MAPKK had no effect on any of the parameters examined. The K97A and S222A mutants served as dominant negatives by suppressing MAPKK, MAPK, and p90rsk activation in vivo. S222E enhanced all of these activities, and S212A,S218A had a small inhibitory effect. A similar trend was observed when cellular proliferation was examined and the different effects were accompanied by altered cellular shape. Taken together, our results demonstrate a direct linkage between the MAPK signaling pathway and the control of cellular proliferation and morphology and also establish that phosphorylation of serine 222 is essential for MAPKK activation together with the phosphorylation of an additional serine(s) (probably serine 218).

3T3 Cells

[Pneumocystis carinii pneumonia in HIV-infected patients examined by bronchoscopy during the period 1989-1991].

We analyzed prospectively collected data from 211 HIV-infected patients who underwent bronchoscopy because of pulmonary symptoms during the period 1989-1991. We found an improvement in survival for patients diagnosed with Pneumocystis carinii pneumonia (PCP) in 1991 as compared to 1989 and 1990 (log rank test: p = 0.07). A significant decline in PO2 and CD4 cell count was observed in patients with PCP in the same period (p < 0.05 and 0.005 respectively). We suggest that the decline in CD4 cell count and in PO2 may reflect a change in Pneumocystis carinii transmission, and that the improved survival may be a result of altered treatment strategy of PCP.

AIDS-Related Opportunistic Infections

Expression of glutamate receptor genes in white matter: developing and adult rat optic nerve.

We have identified the glutamate receptor genes expressed by developing and adult rat optic nerve. Results from PCR suggested that of the ionotropic glutamate receptor (GluR) gene family, only GluR1 and GluR3 subunits are expressed by optic nerve. However, Northern blot analysis demonstrated that only the GluR1 subtype is expressed at appreciable levels. In situ hybridization histochemistry of GluR1 was performed and a diffuse pattern of expression was observed in both postnatal day 15 and adult optic nerve. Restriction mapping of the GluR1 PCR product indicated the expression of the flip alternative splice version in optic nerve. The detected GluR1 message was not due to axons because no loss of expression was observed after the degeneration of axons. PCR analysis also revealed the presence of a metabotropic glutamate receptor in optic nerve.

Aging

Human T-cell mitogen-activated protein kinase kinases are related to yeast signal transduction kinases.

Mitogen-activated protein (MAP) kinase kinases, intermediates in a growth factor-stimulated protein kinase cascade, are dual specificity protein kinases that specifically phosphorylate and activate MAP kinases in response to extracellular signals. Here, we report the cloning of two forms of cDNA that encode this protein from human T-cells. MKK1a encodes a protein with predicted molecular size of 43,439 Da. Overexpression of this clone in COS cells led to elevated levels of protein and phorbol ester-stimulated MAP kinase kinase activity, confirming that MKK1a encodes the predicted protein. MKK1b, which appears to be an alternatively spliced form of the MKK1a gene, encodes a protein with predicted molecular size of 40,745 Da. Northern analysis revealed that the MKK1 cDNA hybridizes with a single 2.6-kilobase mRNA species in all human tissues examined. Sequence comparison shows homology to a group of yeast kinases that participate in signal transduction and to subdomain XI of other dual specificity kinase.

Amino Acid Sequence

Purification and characterization of mitogen-activated protein kinase activator(s) from epidermal growth factor-stimulated A431 cells.

Two peaks of mitogen-activated protein (MAP) kinase activator activity are resolved upon ion exchange chromatography of cytosolic extracts from epidermal growth factor-stimulated A431 cells. Two forms of the activator (1 and 2) have been purified from these peaks, using chromatography on Q-Sepharose, heparin-agarose, hydroxylapatite, ATP-agarose, Sephacryl S-300, Mono S, and Mono Q. The two preparations each contained one major protein band with an apparent molecular mass of 46 or 45 kDa, respectively, on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Evidence identifying the MAP kinase activators as the 46- and 45-kDa proteins is presented. Using inactive mutants of MAP kinase as potential substrates, it was found that each preparation of MAP kinase activator catalyzes phosphorylation of the regulatory residues, threonine 188 and tyrosine 190, of Xenopus MAP kinase. These results support the concept that the MAP kinase activators are protein kinases. These MAP kinase kinases demonstrate an apparent high degree of specificity toward the native conformation of MAP kinase, although slow autophosphorylation on serine, threonine, and tyrosine residues and phosphorylation of myelin basic protein on serine and threonine residues is detected as well.

Animals

Correlates of concern in parents of high-risk infants at age five.

Parents whose children have experienced a high-risk birth may be in double jeopardy due to the biomedical effects of the birth trauma as well as parent perceptions and feelings regarding medical vulnerability and the subsequent impact on caregiving. The present study determined whether the developmental, social, and medical concerns reported by 223 parents of high-risk infants at age 5 were congruent with the child's medical history and developmental performance at age 5. Results indicated that parents with the highest levels of developmental, social, and medical concerns had children with lower birth weights who were born earlier and spent longer periods of time in the hospital at birth. These children also had lower outcomes on the cognitive ability measures at age 5, and were more likely to demonstrate potential behavior problems. Present findings suggest the degree of concern reported by parents of high-risk infants is congruent with the birth history and the developmental status of their child.

Adaptation, Psychological

Differential intracellular calcium responses to glutamate in type 1 and type 2 cultured brain astrocytes.

Fluorescence image analysis using the calcium indicator fluo-3 was used to examine changes in [Ca2+]i induced by glutamate in mixed glia populations cultured from neonatal rat brains. [Ca2+]i responses were correlated with glia type by performing immunohistochemistry using markers specific for type 1 and type 2 astrocytes on the same cells used in the imaging experiments. Glutamate (30-500 microM) induced two markedly different [Ca2+]i responses in the two astrocyte types: the response in type 1 astrocytes consisted of an initial fast transient followed by varying degrees of oscillations, whereas the predominant response in type 2 astrocytes was a slow rise in [Ca2+]i to a more or less sustained and nonoscillatory level. In some type 2 astrocytes, an initial spikelike transient similar to that in type 1 astrocytes was observed; the overall size of the spike, however, was smaller than in type 1 astrocytes. Two agonists for the ionotropic glutamate receptor, alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA) and kainate, elicited a 6-cyano-7-dinitroquinoxaline-2,3-dione (CNQX)-sensitive, external Ca(2+)-dependent, sustained [Ca2+]i rise in type 2 but not type 1 astrocytes. The initial spike in type 2 astrocytes was less dependent on external Ca2+ and not blocked by CNQX. [Na+]i as measured by the Na(+)-fluorescence dye SBFI, was elevated by kainate in both astrocyte types, though the increase was larger in type 2 astrocytes. This increase was reduced by CNQX, suggesting this [Ca2+]i increase was mediated, at least in part, by ionotropic glutamate receptors. The results are discussed in terms of the relative distribution of two classes of glutamate receptors on these two astrocyte types: one, the ionotropic class, is linked directly to an ion channel, and the other, the metabotropic class, induces internal mobilization of Ca2+ via inositol phospholipid hydrolysis.

6-Cyano-7-nitroquinoxaline-2,3-dione

Global assessment of patients--a bedside study. II. Inter-observer variation and frequency of clinical findings.

Three physicians independently assessed 201 hospital in-patients for the presence of 10 basic physical characteristics, and made an overall assessment of whether or not the patients appeared ill. There were significant differences between the physicians in the average number of observations recorded for each patient (P less than 0.001). There was a significant difference (P less than 0.05) between the physicians regarding the number of positive diagnoses of anaemia, abnormal nutritional state, breathing difficulties and the overall assessment. Inter-observer variation was estimated by a pairwise comparison of the three physicians. Overall agreement rates ranged from 0.65 for abnormal nutritional state to 0.99 for presence of pain. After adjusting for random agreement, kappa values between 0.09 (elevated body temperature) and 0.88 (consciousness impaired) were found. No tendency towards a higher level of agreement in the overall assessment than in the basic findings was observed.

Diagnosis

Epidemiology of bovine virus diarrhoea virus.

A better understanding of the epidemiology and pathogenesis of bovine virus diarrhoea virus (BVDV) has emerged in recent years. Fetal infections and in particular those resulting in birth of persistently infected calves are of central importance for the epidemiology of BVDV. A prevalence of persistently infected, viraemic animals of about 1% is found in Denmark and elsewhere by examination of randomly collected blood samples. A recent field study shows that 53% of randomly selected herds in an area in Denmark where BVDV is endemic had one or more persistently infected animals. Persistently infected cows may breed and will always transmit the infection to the calf. Such familial occurrence of persistent infection seems to be a fairly common phenomenon. Persistently infected cattle are important sources of infection to other cattle. Transiently infected cattle following experimental exposure will usually not transmit the infection by contact but this may not always apply to cattle after natural infection. Knowledge of the occurrence and potential for spread of virus from persistently infected bulls is reviewed. Virus is excreted with semen of both persistently and transiently infected bulls and BVDV may be transmitted by use of infected semen for insemination. The potential for spread of the infection through embryo transfer should be avoided by the use of adequate testing and controls.

Animals

Fluorescence measurement of changes in intracellular calcium induced by excitatory amino acids in cultured cortical astrocytes.

Population response of [Ca2+]i in cultured cortical astrocytes to excitatory amino acids was measured at room temperature using the calcium-sensitive dye fura-2. Quisqualic acid (QA), glutamate (Glu), and kainic acid (KA) caused a peak increase in [Ca2+]i in the order QA greater than Glu greater than KA. No response to N-methyl-D-aspartic acid (NMDA) was observed whether or not Mg2+ was present externally. Both QA and Glu (100 microM) frequently elicited a decaying oscillatory [Ca2+]i response during sustained agonist application; the period of oscillations initially was 23.5 sec and increased as the response was damped. Comparatively, the [Ca2+]i response to KA was nonoscillatory. Both responses to Glu and KA were reduced slightly by antagonist gamma-D-glutamylaminomethyl-sulfonic acid (1 mM), but virtually were abolished by kynurenic acid (3 mM). Replacement of external Na+ by choline had no significant effect on the Glu response. Removal of external Ca2+ reduced the peak response to QA, Glu, and KA to 40, 34, and 18%, respectively; and markedly reduced the degree of QA- and Glu-induced [Ca2+]i oscillations. Pretreatment with phorbol esters, a potent activator of protein kinase C, blocked the [Ca2+]i response to Glu but not KA. It is concluded that cortical astrocytes express Glu receptors of the non-NMDA type in culture and that receptor activation leads to Ca2+ influx and release of internal Ca2+. Mobilization of Ca2+ apparently occurs via the known Glu-mediated hydrolysis of inositol lipids, which may come under negative-feed-back control by protein kinase C activation. Oscillatory [Ca2+]i signaling offers the possibility of a dynamic population response in an electrically coupled glial network.

Amino Acids