Biomedical subjects

M Kao

Publications and source records attributed to M Kao.

At least 19 recordsLinked to original sources

PubMed2001

Validation of the sterilization procedure of allogeneic avital bone transplants using peracetic acid-ethanol.

Different procedures are available to inactivate bacteria and fungi, including their spores, as well as viruses in human bone transplants. The most efficient methods are considered to be gamma irradiation and thermal inactivation as well as chemical sterilization methods like the peracetic acid-ethanol treatment (PES). Following national and international standards or draft standards, the antimicrobial effectiveness of this procedure was evaluated. Due to the standardizable size as well as the clinical relevance, defatted human spongiosa cuboids (15x15x15 mm) served as model system. After treatment with PES for 2 and 4 hours, respectively, the titre of living micro-organisms was determined in the supernatant and the cuboid. A reduction in the titre of viable micro-organisms below the detection level (reduction factor >5 log10) was already achieved after an incubation time of 2 hours (Staphylococcus aureus, Enterococcus faecium, Pseudomonas aeruginosa, Bacillus subtilis, Clostridium sporogenes, Mycobacterium terrae, Candida albicans as well as spores of Bacillus subtilis). No viable micro-organisms could be detected in any of the PES-treated test cuboids. Spores of Aspergillus niger were also completely inactivated. The PES procedure proved to be a reliable method for the sterilization of human bone transplants derived from spongiosa.

Bacteria↗

PubMed2000

Elevated topoisomerase I activity in cervical cancer as a target for chemoradiation therapy.

OBJECTIVE: The aim of this study was to determine whether the activity of topoisomerase I (topo I), the target of the anti-neoplastic drug camptothecin (CPT), is elevated in cervical cancer and whether CPT can radiosensitize cervical tumors. METHODS: The topo I activity of 11 normal cervix and 30 cervical carcinoma tumors was assayed by measuring the relaxation of supercoiled DNA. Subconfluent or postconfluent CaSki human cervical carcinoma cells were exposed to CPT (1-5000 ng/ml) and immediately X-irradiated (0-800 cGy). Cell survival was determined by clonogenic assay. RESULTS: Mean topo I activity in cervical cancer (3.0 +/- 0.06 h(-1)) was significantly greater than in normal cervix tissue (0.29 +/- 0.06 h(-1)). Stage 3 and 4 cervical carcinoma specimens displayed a trend of greater topo I activity (5.88 +/- 3.7 h(-1)) than stage 1 and 2 tumors (2.57 +/- 0.47 h(-1)). No correlation between topo I protein levels and catalytic activity was found. Combined treatment of subconfluent CaSki cells with CPT and ionizing radiation resulted in additive killing of cells. Combined treatment of postconfluent CaSki cells with low doses of radiation (200 and 400 cGy) and 1 or 10 ng/ml CPT for 2 or 48 h produced significant cytotoxicity compared to CPT or radiation alone, which were ineffective at these doses. CONCLUSIONS: Topo I activity is elevated in cervical cancer compared to normal cervix. The radiosensitivity of noncycling cells within cervical tumors may be increased by simultaneous treatment with low doses of CPT or other topo I inhibitors.

Antineoplastic Agents, Phytogenic↗

PubMed2000

Further studies on thermal resistance of bovine parvovirus against moist and dry heat.

To supplement the results of thermal resistance of bovine parvovirus (Haden strain, BPV) published previously, we carried out assays at 60 degrees C (moist heat) to compare the thermal resistance of BPV with that of hepatitis B-virus (HBV). What we know about the resistance of HBV at a temperature of 60 degrees C is mainly based on data collected within the context of blood product pasteurization. The results suggest that at a temperature of 60 degrees C, BPV shows thermal resistance comparable to HBV. Thus, BPV--which is easier to handle--can be considered a good test virus to verify the efficacy of thermal disinfection techniques against HBV. BPV is very resistant against dry heat of 100 degrees C, the inactivation largely depending upon the residual moisture of the lyophilisate. Reducing the residual moisture from 2% to less than 1%, the exposure time has to be prolonged by ca. 2.5 times to achieve the same virucidal effect.

Animals↗

PubMed1999

Virus safety of avital bone tissue transplants: evaluation of sterilization steps of spongiosa cuboids using a peracetic acid-methanol mixture.

The aim of this study was to validate the virus-inactivating/eliminating capacity of the manufacturing process of spongiosa cuboids. Both the sterilization step with peracetic acid (PAA)/ethanol and the defatting step of bones with chloroform/methanol (2:1, v/v) were investigated. Relevant enveloped, non-enveloped, and model viruses belonging to different virus families were included in the investigation: human immunodeficiency virus type 2 (HIV-2), hepatitis A virus (HAV), poliovirus (PV-1), pseudorabies virus (PRV), porcine parvovirus (PPV), and bovine virus diarrhoea virus (BVDV). Treatment of virus-spiked spongiosa cuboids for 4 hours at room temperature (RT) with 1% PAA/24% ethanol (PES) efficiently inactivated most viruses. Titres were reduced by more than 4 log(10)with the exception of HAV. The defatting step with chloroform/methanol reduced HAV titres by a factor of >/=7.0 log(10). From these results it can be concluded that the treatment of spongiosa cuboids with (i) chloroform/methanol and (ii) 1% PAA/24% ethanol solution leads to a virus-safe medicinal product.

Bone Transplantation↗

PubMed1995

Herpesvirus-like infection in a raccoon (Procyon lotor).

During March 1990, a subadult raccoon found dead in northeastern Pennsylvania (USA) had gross lesions of multifocal hepatitis. Microscopically, multifocal randomly distributed areas of acute necrosis with intranuclear viral inclusions were seen in liver, spleen, adrenal glands, and tongue. Ultrastructural and immunoperoxidase results of formalin fixed liver were compatible with herpesvirus infection. This virus could be unique to the raccoon or may have been acquired from another species.

Animals↗

PubMed1993

A novel adenylyl cyclase sequence cloned from the human erythroleukemia cell line.

The polymerase chain reaction (PCR) was used to detect several forms of adenylyl cyclase (AC's) expressed in human erythroleukemia (HEL) cells. Degenerate oligonucleotide primers were synthesized based on the conserved sequences in the C2a area of the AC's. HEL cells were found to contain mRNA for type III and type VI AC. In addition, a novel AC message was identified. The cloned sequence, excluding primer areas, represented 69 amino acids with most similarity to rat AC's II and IV. Northern analysis of RNA from HEL cells demonstrated a 6.7 kilobase message. RNase protection assays revealed that in HEL cells the novel AC mRNA was dominant compared to types III and VI. Human embryonic kidney cells (HEK293) were also used a source of mRNA to synthesize cDNA for PCR reactions. The HEK293 cells were found to contain message corresponding to type II, III, VI AC as well as the novel AC message. The novel AC message was also detected in human brain tissue and was most abundant in the caudate, cerebellum and hippocampus. The smallest amount of novel AC mRNA in the tested brain tissue was found in the cortex. The mRNA for the novel AC was relatively abundant in human liver.

Adenylyl Cyclases↗

PubMed1993

Detection of antibodies against Borna disease virus in sera and cerebrospinal fluid of horses in the USA.

Sera from 295 horses in the USA were examined by an indirect immunofluorescence assay and Western blot assays to determine the prevalence of Borna disease virus infection. Eight (2.7 per cent) of the samples were positive in both assays, and 18 (6.1 per cent) were positive only in the Western blot assay. The indirect fluorescence titres ranged from 1:20 to 1:80 of antibodies recognising the virus-specific antigen from Borna disease virus-infected cells. The purified virus-specific proteins isolated from infected rat brains were recognised by positive equine serum samples after immunostaining by a Western blot technique. Information obtained from the owners about the history of the seropositive horses revealed that they were either clinically normal or had a pathological diagnosis of disease unrelated to Borna disease. This is the first report of the detection of antibodies to Borna disease virus in horses in the USA. The disease may be more widespread in a subclinical form, with very long incubation periods, and may not necessarily be restricted to historically endemic areas.

Animals↗

PubMed1993

Detection of Borna disease virus-reactive antibodies from patients with affective disorders by western immunoblot technique.

Borna disease (BD) virus is a partially characterized neurotropic agent with a predilection for neurons and astrocytes in the limbic system and cerebrum of infected hosts. Although it usually causes a fatal encephalitis, some laboratory animals which have been experimentally inoculated can develop a persistent non-fatal infection characterized by a neuro-behavioral syndrome akin to human manic-depression. Using immunofluorescent techniques, we previously observed BD virus-specific antibodies in the sera of 4.5% of affectively ill patients, with the highest titers present in bipolar patients. More recently, we have developed a sensitive Western blot assay for the detection of anti-BD virus antibodies to a 38/40 kDa and 24 kDa protein in human serum. In the present study, we screened 138 affectively ill patients and 117 healthy controls and observed a significantly great proportion of patients with antibodies to the 38/40 kDa protein (P < 0.0001), the 24 kDa protein (P < 0.05) and both the 38/40 kDa and 24 kDa proteins (P < 0.025). These data extend prior reports on the presence of BD virus-specific antibodies in psychiatric patients, and suggest that a BD virus-like agent may be associated with affective illness in humans.

Adult↗

PubMed1993

Rabies and borna disease. A comparative pathogenetic study of two neurovirulent agents.

BACKGROUND: Rabies and Borna disease viruses have been regarded as classical neurotropic agents. Many pathogenetic similarities are shared by these two negative strand RNA viruses. In view of recently gained data on the virology and pathology of these two diseases, and up-to-date comparative pathogenetic study seems to be justified. EXPERIMENTAL DESIGN: This study is based on a survey of experimental and natural infections of laboratory animals and natural hosts. The morphologic damage to the nervous system has been evaluated by light and electron microscopy, with special emphasis on immunocytochemical methods. RESULTS: This comparative study disclosed that both viruses are transported inside axons, pass synapses and propagate along neuronal networks. At the sites of synaptic transfer, full virus particles can never be detected in the early phase of rabies virus infection; in Borna disease virus (BDV) infection, virus particles cannot be found in any phase of disease progression. Thus, a major difference exists between the two agents insofar as rabies virus is morphologically well characterized, whereas BDV has never been visualized in tissue sections. Furthermore, rabies virus infects only neurons, whereas BDV also infects glial cells. The host range and the scale of infection of extraneural tissues by both agents is extremely similar. CONCLUSIONS: These observations allow us to postulate that the synaptic transfer of both viruses likely ensures in the form of bare nucleocapsids (ribonucleoprotein-transcriptase complexes). While in the later phases of replication complete rabies virions are regularly assembled, BDV propagates within the central nervous system in an incomplete form, so that it remains morphologically imperceptible. Thus, BDV may appear in a complete, enveloped form only when exiting the host organism. The dissemination patterns of the two agents may be influenced by specific affinities to neurotransmitter receptor sites. It remains unresolved, why BDV readily infects non-neuronal central nervous system cells, while rabies virus remains restricted to neuronal elements.

Animals↗

PubMed1992

Delineation of putative mechanisms involved in antibody-mediated clearance of rabies virus from the central nervous system.

The in vitro biological activities of several rabies virus-neutralizing monoclonal antibodies (mAbs) were compared with their ability to prevent a lethal rabies virus encephalomyelitis. The protective activity of a particular mAb in vivo did not correlate with its virus-neutralizing activity in vitro; rather it was related to the mAb's ability to inhibit virus spread from cell to cell and to restrict rabies virus RNA transcription. Since treatment of rabies virus-infected cells with virus-neutralizing mAbs results in an endocytosis of the antibody, we hypothesize that an antibody may exert its inhibitory activity even after uptake by the cell. Post-exposure treatment of rats with a mAb that inhibited both virus spread and virus RNA transcription in vitro resulted in viral clearance from the central nervous system and protected the animals against a lethal rabies virus infection.

Actins↗

PubMed1992

Kinetics of virus spread and changes in levels of several cytokine mRNAs in the brain after intranasal infection of rats with Borna disease virus.

We have used the reverse transcriptase-polymerase chain reaction technique to gain insight into the pathogenesis of encephalitis caused by Borna disease virus (BDV). RNA specific for BDV was first detected in the olfactory bulb of intranasally infected rats at 6 days postinfection (p.i.). At 14 days p.i., high levels of BDV RNA were found in all brain regions, and at 26 days p.i., BDV-specific RNA was also present in the eye, nasal mucosa, and facial skin. In the chronic phase of the disease, BDV RNA was identified in many peripheral organs but not in blood. Analysis of brain tissue for the presence of cytokine mRNAs revealed that the mRNA levels of interleukin-6 (IL-6), tumor necrosis factor alpha, and IL-1 alpha had increased sharply at 14 and 26 days p.i. These cytokine mRNAs reached maximum levels at the peak of inflammatory reactions and decreased drastically in the chronic phase of the disease. Although IL-2 mRNA was also found in normal brain, it was markedly increased in BDV-infected brain at 14 days p.i. Expression of gamma interferon (IFN-gamma) mRNA, which was not observed in normal rat brain, was detected at 14 days p.i. and reached a maximum level at 38 days p.i. IL-2 and IFN-gamma mRNA expression correlated with expression of CD4 and CD8 mRNAs, indicating that both CD4+ and CD8+ T lymphocytes are induced in the early stages of BDV infection. Since IFN-gamma and CD8 mRNA levels were still highly elevated in the chronic phase of Borna disease, it is likely that CD8+ T lymphocytes act to reduce inflammation and to ameliorate neurological signs during the chronic phase of infection.

Animals↗

PubMed1991

Neuronal autophagy in experimental scrapie.

In this study we report the formation of giant autophagic vacuoles (AV) in neurons in experimental scrapie in hamsters. Autophagy is an important step in the cellular turnover of proteins and organelles. It is known to occur in neurons under physiological as under pathological conditions. Giant AV, however, are seen very rarely only in pathological states. In our model AV are much more numerous after intracerebral (i.c.) transmission of the scrapie agent than after the transmission via the intraperitoneal route which points to a correlation between the intensity of the process and the period of incubation. As the appearance of the AV in our model is correlated chronologically with that of scrapie-associated fibrils, at least after i.c. transmission, the process may be related to a disturbance of cellular protein metabolism and, thus, to the processing of prion protein.

Animals↗

PubMed1990

Technique for breast irradiation using custom blocks conforming to the chest wall contour.

A technique for the treatment of the breast and regional nodes is presented. The technique involves the use of tangential fields to treat the breast and chest wall. Customized blocks which conform to the slope of the chest wall are made for each tangent field. Simulation and treatment with this technique requires no special equipment. The setup is simple and quick. A three-field technique is also described using the custom half-beam blocks; this technique avoids the use of tangential field corner blocks, thus simplifying simulation and treatment.

Breast↗

PubMed1990

Transcriptional regulation of the murine TSH subunit genes.

These studies have demonstrated that the murine TSH subunit genes are sensitively regulated by thyroid hormone. The T3 receptor complex interacts with both the TSH beta and alpha-subunit gene either in or near the 5' flanking region in close proximity to the transcriptional start sites. This interaction interrupts transcription from the start sites, thus decreasing transcription of the two genes. As a result steady state mRNA levels of both TSH beta and alpha-subunit genes are decreased in the cytoplasm of thyrotropic cells. This series of interactions explains most of the effects of T3 on TSH biosynthesis.

Animals↗

PubMed1989

Learning deficiencies in Borna disease virus-infected but clinically healthy rats.

Borna disease (BD) virus, a still unclassified neurotropic agent, causes either fatal encephalomyelitis or persistent asymptomatic infection in a variety of animal species. We monitored the neuronal functions of intracerebrally infected but healthy rats with three types of learning experiments. Spatial discrimination learning, using the y maze and the hole board, was significantly less successful in BD virus-infected (I) compared with mock-infected (M) rats. Similarly, I rats tended to show a certain emotional disturbance (reduced resting behavior and less anxiety) as evaluated by open-field and neophobia tests. Furthermore, in two aversive learning experiments (taste aversion and reaction suppression via Skinner box), it appeared that the I rats expressed a significantly diminished ability to learn pain avoidance compared with M rats. In conclusion, we found specific learning deficiencies together with subtle behavioral alterations suggesting that BD virus causes certain modulations of high integrative brain functions which are only detectable under experimental conditions.

Animals↗

PubMed1988

Possible role of cytosolic free calcium concentrations in mediating insulin resistance of obesity and hyperinsulinemia.

Insulin- and glyburide-stimulated changes in cytosolic free calcium concentrations [( Ca2+]i) were studied in gluteal adipocytes obtained from six obese women (139 +/- 3% ideal body wt) and six healthy, normal weight age- and sex-matched controls. Biopsies were performed after an overnight fast and twice (at 3 and 6 h) during an insulin infusion (40 mU/m2 per min) (euglycemic clamp). In adipocytes obtained from normal subjects before insulin infusion, insulin (10 ng/ml) increased [Ca2+]i from 146 +/- 26 nM to 391 +/- 66 nM. Similar increases were evoked by 2 microM glyburide (329 +/- 41 nM). After 3 h of insulin infusion, basal [Ca2+]i rose to 234 +/- 21 nM, but the responses to insulin and glyburide were completely abolished. In vitro insulin-stimulated 2-deoxyglucose uptake was reduced by insulin and glucose infusion (25% stimulation before infusion, 5.4% at 3 h, and 0.85% at 6 h of infusion). In obese patients, basal adipocyte [Ca2+]i was increased (203 +/- 14 nM, P less than 0.05 vs. normals). The [Ca2+]i response demonstrated resistance to insulin (230 +/- 23 nM) and glyburide (249 +/- 19 nM) stimulation. Continuous insulin infusion increased basal [Ca2+]i (244 +/- 24 nM) and there was no response to either insulin or glyburide at 3 and 6 h of study. Rat adipocytes were preincubated with 1-10 mM glucose and 10 ng/ml insulin for 24 h. Measurements of 2-deoxyglucose uptake demonstrated insulin resistance in these cells. Under these experimental conditions, increased levels of [Ca2+]i that were no longer responsive to insulin were demonstrated. Verapamil in the preincubation medium prevented the development of insulin resistance.

Adipose Tissue↗

PubMed1988

Relationship between cytosolic free calcium concentration and 2-deoxyglucose uptake in adipocytes isolated from 2- and 12-month-old rats.

We have examined the relationship between insulin-stimulated 2-deoxyglucose uptake and cytosolic free calcium concentrations, [( Ca2+]i), in adipocytes isolated from 2- and 12-month-old rats. The basal rates of glucose uptake and the levels of cytosolic Ca2+ were only minimally reduced in 12-month-old animals. In contrast, insulin-stimulated glucose up-take and [Ca2+]i were significantly decreased in older adipocytes at all insulin concentrations (P less than 0.01). When the rate of glucose uptake was plotted as a function of [Ca2+]i, insulin-stimulated glucose uptake was almost identical in older and younger animals at any given level of [Ca2+]i. Similar to insulin, glyburide and K+ increased [Ca2+]i in both younger and older adipocytes. However, glyburide- and K+-elicited responses were lower in older rats (P less than 0.01). The effects of insulin, glyburide, and K+ on [Ca2+]i are mediated via voltage-dependent Ca2+ channels. Thus, the present observations suggest an impairment in either function and/or availability of the voltage-dependent Ca2+ channels in older animals. This was supported by the finding of reduced [3H]nitrendipine binding in adipocytes isolated from older animals (6.5% vs. 3.3% in 2- and 12-month-old rats, respectively; P less than 0.01). The results of these experiments indicate that the postreceptor changes in adipocyte responsiveness to insulin in aging may involve inadequate increases in [Ca2+]i. The latter probably occurs as a result of decreased availability and/or function of the voltage-dependent calcium channels.

Adipose Tissue↗

PubMed1987

The existence of an optimal range of cytosolic free calcium for insulin-stimulated glucose transport in rat adipocytes.

We have examined the effects of extracellular and intracellular Ca2+ concentrations upon basal and insulin-stimulated 2-deoxyglucose uptake in isolated rat adipocytes. In the absence of extracellular Ca2+, both basal and insulin-stimulated glucose uptake were significantly reduced. Insulin-stimulated glucose transport was optimal at 1 and 2 mM Ca2+. Further increases in extracellular Ca2+ concentration (3 mM) significantly diminished insulin-stimulated glucose uptake. When intracellular Ca2+ concentrations were augmented by ionomycin (1 microM), insulin-stimulated glucose uptake was significantly reduced at extracellular Ca2+ concentrations of 2 and 3 mM. The levels of intracellular free Ca2+ concentrations were then measured with Ca2+ indicator fura-2. The correlation between the levels of intracellular free Ca2+ and the magnitude of insulin-stimulated glucose uptake revealed that the optimal effect of insulin is observed at Ca2+ levels between 140 and 370 nM. At both extremes outside of this window, both low and high levels of intracellular Ca2+ result in diminished cellular responsiveness to insulin. These data suggest that intracellular calcium concentrations may exert a dual role in the regulation of cellular sensitivity to insulin. First, there must exist a minimal concentration of intracellular calcium to promote insulin action. Second, increased levels of intracellular calcium may provide a critical signal for diminution of insulin action.

Adipose Tissue↗