Biomedical subjects
Publications and source records attributed to J Zhao.
The present paper summarises our recent studies on buffalo babesiosis in the Hubei province of China. It describes the pathogen and the epidemiology of the disease, the tick vector and its biology, control, immunological diagnosis, methods of in vitro cultivation of the pathogen and their practical applications. The identify of the pathogen is discussed in detail.
This paper summarizes a recent study on the in vitro cultivation of Babesia from buffaloes. Buffalo Babesia parasites were cultured many times by the microaerophilous stationary phase (MASP) method of Levy and Ristic (1980) with some modifications. The culture-derived Babesia organisms were similar to forms seen in the blood of infected buffaloes and the pathogenicity of these organisms was unchanged. Factors influencing the in vitro cultivation are discussed in detail.
Packed erythrocytes infected with Babesia bovis were mixed with an equal volume of 16% dimethyl sulphoxide (DMSO) in Alsever's solution and dispensed into 1.5 or 5 ml cryotubes. The vials were kept in liquid nitrogen (-196 degrees C) for 26, 78, 142 or 149 days. The samples were removed from the liquid nitrogen container and rapidly thawed in a 40 degrees C water bath. The thawed blood successfully infected splenectomised buffalo calves by injection via subcutaneous or intravenous or via intravenous and subcutaneous routes. The parasites, typical B. bovis, were discovered in red blood cells 5, 8 or 9 days after inoculation. The highest percentage of parasitised erythrocytes (PPE) was 15%. The babesiosis resulting from cryopreserved parasites was the same as that resulting from fresh parasites inoculated by ticks. Typical clinical signs were found, such as continuous fever (the highest temperature was 41.3 degrees C), anaemia, icterus and haemoglobinuria. Infected calves, which were not treated, died. Cryopreservation is a simple and reliable method for longterm preservation of B. bovis of water buffaloes.
Theileria annulata infection (TAI) is one of the most serious diseases of cattle in Xinjiang Autonomous Region of Uigur Minority Nationality. It has been recorded in 14 prefectures, except the Tulufan Prefecture, but the enzootic areas are mainly distributed around the Zhunger Basin and Talim Basin. According to the records collected in the ten years before vaccination with the schizont cell culture vaccine was carried out, the average incidence rate of TAI in enzootic areas was 7.22% and the mortality rate was 24%. The milk production of cattle suffering from TAI was sharply decreased, and there were usually abortions in pregnant cows. The incidence rate and mortality rate were greater in high grade cattle, so TAI was a constraint to improving cattle breeds. To control this disease effectively in Xinjiang, researchers at the Xinjiang Academy of Animal Science began to study the schizont cell culture vaccine in 1972. In 1977 an immortalised cell line was achieved from a primary cell culture starting with white blood cells from cattle suffering from acute TAI caused by an artificial tick bite. The cell culture medium mainly consisted of calf serum, lactalbumin-hydrolysate, Eagles' medium DMEM and three antibiotics. As a vaccine, the above cells were mixed with preserving medium containing gelatin. This paper describes the experiments on the immunological properties of the vaccine carried out in subsequent years. Up to 1996, vaccine doses for 1,186,150 cattle have been produced and sold. This vaccine has had a critical effect on the control of TAI in Xinjiang. Owing to the sharp decrease in the incidence rate and mortality rate of TAI after cattle were vaccinated, the annual economic benefit of the vaccine is at least 1,620,000 yuan.
The ryanodine receptor (RyR) of skeletal muscle contains two functional domains: a carboxyl-terminal hydrophobic domain that forms the putative conduction pore of the calcium release channel, and a large cytoplasmic domain that corresponds to the "foot structure." To understand the contribution of the foot structure to the function of the calcium release channel, we studied a RyR deletion mutant, delta(1641-2437)-RyR, in which a region that is rich in glutamate and aspartate residues (a.a. 1641-2437) was removed. The wild-type and delta(1641-2437)-RyR proteins were expressed in a Chinese hamster ovary (CHO) cell line, and functions of single calcium release channels were measured in the lipid bilayer membrane. The wild-type RyR forms functional calcium release channels with a linear current-voltage relationship similar to that of the native channel identified in the sarcoplasmic reticulum membrane of skeletal muscle, whereas the channels formed by delta(1641-2437)-RyR exhibit significant inward rectification, i.e., currents moving from cytoplasm into SR lumen were approximately 20% less than that in the opposite direction. As in to the wt-RyR channel, opening of the delta(1641-2437)-RyR channel has a bell-shaped dependence on the cytoplasmic calcium, but the calcium-dependent activation and inactivation processes of the delta(1641-2437)-RyR channel are shifted to higher calcium concentrations. Our data show that deletion of a.a. 1641-2437 from the foot region of the skeletal muscle RyR results in changes in both ion conduction and calcium-dependent regulation of the calcium release channel.
The ryanodine receptor (RyR) is one of the key proteins involved in excitation-contraction (E-C) coupling in skeletal muscle, where it functions as a Ca2+ release channel in the sarcoplasmic reticulum (SR) membrane. RyR consists of a single polypeptide of approximately 560 kDa normally arranged in a homotetrameric structure, which contains a carboxyl (C)-terminal transmembrane domain and a large amino (N)-terminal cytoplasmic domain. To test whether the carboxyl-terminal portion of RyR is sufficient to form a Ca2+ release channel, we expressed the full-length (RyR-wt) and C-terminal (RyR-C, approximately 130 kDa) RyR proteins in a Chinese hamster ovary (CHO) cell line, and measured their Ca2+ release channel functions in planar lipid bilayer membranes. The single-channel properties of RyR-wt were found to be similar to those of RyR from skeletal muscle SR. The RyR-C protein forms a cation-selective channel that shares some of the channel properties with RyR-wt, including activation by cytoplasmic Ca2+ and regulation by ryanodine. Unlike RyR-wt, which exhibits a linear current-voltage relationship and inactivates at millimolar Ca2+, the channels formed by RyR-C display significant inward rectification and fail to close at high cytoplasmic Ca2+. Our results show that the C-terminal portion of RyR contains structures sufficient to form a functional Ca2+ release channel, but the N-terminal portion of RyR also affects the ion-conduction and calcium-dependent regulation of the Ca2+ release channel.
Within a homogeneous population of cells, the tendency of individual cells to become diseased is not 'homogeneous'. As a general concept of disease, the 'liability theory' describes such a tendency (cell liability) as a quantitative characteristic following a continuous distribution. This suggests that there are always some cells that are more liable than the rest and that the liability values of cells or liability-associated cell characteristics can be assessed quantitatively. By defining the more or extremely liable cells through a critical value or 'threshold', the liability theory hypothesizes that these cells may play a significant role in determining or affecting the overall disease liability of a cell population; the more such cells are present, the more liable the cell population is, as a whole, to disease. Furthermore, it proposes that the origin and development of disease may be fundamentally relevant to this kind of intrapopulational variation in cell liability. The liability theory attempts to provide a theoretical framework for studying the 'cell population' mechanism of disease, or cell population pathology. Also, it may serve as a working model for analysis and assessment of cell population liabilities for various diseases.
A case of superior dislocation of the mandibular condyle into the middle cranial fossa is presented. A modified method of reducing the mandibular condyle was used, and restitution of the perforated glenoid fossa was achieved by a combination of titanium plate and repair of the articular disk.
It is known that intracellular pH drops rapidly after the onset of ischemia in cardiac muscle and may play some role in the rapid drop in force that ensues. It is also known that alpha 1-adrenoceptor agonists alkalinize intracellular pH by stimulating Na+/H+ exchange and may represent a mechanism which facilitates recovery of intracellular pH from acidosis. Lowering or raising pH shifts the Ca2+ dependence of force development in muscle fibres to higher or lower free Ca2+ concentrations, respectively, yet the precise mechanism is unknown. To investigate this phenomenon we have used skinned skeletal or cardiac muscle fibres whose endogenous troponin C (TnC) has been replaced with chicken skeletal TnC labelled with DANZ (STnCDANZ) or recombinant cardiac TnC labelled with IAANS (CTnC3(C84)[AANS), respectively. The fluorescence of the STnCDANZ or CTnC3(C84)IAANS was enhanced by Ca2+ binding to the Ca(2+)-specific (regulatory) site(s) of STnC or CTnC when incorporated into skinned fibres, and was measured simultaneously with force. When the pH was changed from 7.0 to 6.5 or 7.5 the shift in the Ca2+ dependence of force paralleled the shift in fluorescence. Since the force and fluorescence shift in parallel as the pH is lowered or raised, it can be concluded that these changes in Ca2+ sensitivity are caused by a decrease or increase, respectively, in the Ca2+ affinity of the Ca(2+)-specific site(s) of TnC. Since lowering or raising the pH also resulted in lower or higher, respectively, maximal Ca2+ activated force while maximal fluorescence remained unchanged, it is possible that H+ may act indirectly, as well, by reducing or increasing, respectively, the number or type of crossbridges attached to actin and thereby alter the crossbridge induced depression or elevation, respectively of the observed TnC Ca2+ affinity. Experiments with 2,3-butanedione monoxime, however, where force-generating crossbridges were greatly reduced, indicated that the pH effect may be primarily related to a direct change in the Ca2+ affinity to the regulatory sites of TnC.
Expression of opioid receptor-like receptor (ORL1) and its endogenous peptide agonist nociceptin/orphanin FQ (N/OFQ) during mouse embryogenesis have been investigated. Transcripts of ORL1 and N/OFQ were detected by RT-PCR in mouse brain of day 8 embryo (E8) and the expression continued afterwards. Northern blot analysis revealed abundant expression of ORL1 at postnatal day 1 (P1) and N/OFQ at E17 and P1 in the brain but none was detected in other embryonic tissues. The presence of functional ORL1 in mouse embryonic brain was also confirmed by specific binding of [3H] N/OFQ (kd = 1.3 +/- 0.5 nM and Bmax = 72 +/- 9 fmol/mg protein) as well as by N/OFQ-stimulated G protein activation.
The opioid receptor-like1 receptor (ORL1), an orphan receptor whose human and murine complementary DNAs, has been characterized recently. ORL1 transcripts are particularly abundant in the central nervous system. We demonstrated that ORL1 expressed in human neuroblastoma SK-N-SH and SH-SY5Y cell lines by radioligand binding assay, reverse transcription polymerase chain reaction (RT-PCR) and Northern analysis in the present study. Stimulation with ORL1 specific agonist, nociceptin/orphanin FQ, increased [35S]GTP gamma S binding to SK-N-SH cell membranes (EC50 = 14 +/- 0.45 nM), and attenuated forskolin-stimulated accumulation of cellular cAMP (EC50 = 0.80 +/- 0.45 nM), indicative that activation of ORL1 activates G proteins and inhibits adenylyl cyclase. Activation of ORL1 receptor was also accessed using CHO:hORL1 cell line by microphysiometer. Treatment of nociceptin/orphanin FQ increased extracellular acidification rate significantly.
Replication-defective (E1-E3-deleted) human adenovirus vectors are a promising means of therapeutic gene delivery to skeletal muscle cells. Since the tropism of adenovirus is nonselective, muscle-specific expression of systemically administered vectors can only be achieved by the use of a tissue-specific promoter/enhancer that is small enough to fit the insert capacity of the vector. We have generated two replication-defective adenovirus recombinants (AV) in which the reporter gene (either firefly luciferase or E. coli beta-galactosidase) was driven by a truncated (1.35 kb) muscle creatine kinase (MCK) promoter/enhancer or by the fast troponin I (TnI) promoter/enhancer. Highly efficient and muscle-specific transgene expression was demonstrated in immunodeficient mice after local injection of AV into muscles at an early age. In nonmuscle tissues (brain, liver, kidney, lung), the transgene expression was extremely low even though in these tissues in situ polymerase chain reaction showed as high an infectivity of the cells by the AV as in muscle. The relatively small size, the good efficiency and the muscle specificity of the MCK promoter would make it ideal to drive the 6.3 kb (truncated) dystrophin cDNA in first generation AV (with a limited (8 kb) insert capacity) designed for gene therapy of Duchenne muscular dystrophy.
Although social and cultural influences are clearly important, family, twin and adoption studies indicate that genes contribute significantly to substance abuse. Substance abuse is associated with novelty seeking, a heritable human personality trait which may be influenced by alleles of the dopamine D4 (DRD4) gene exon III VNTR. Consequently Kotler et al analysed the DRD4 VNTR in opiate-dependent subjects from Israel, and found a significant excess of the 7-repeat allele. We have attempted to replicate this finding using a Han Chinese case-control sample of 121 heroin-dependent subjects and 154 normal controls. We found two 7-repeat alleles which occurred exclusively in the patient group, and overall there was an excess of longer alleles, which did not reach significance (chi 2 = 7.04; P = 0.07). When the D4 VNTR was divided into 'long' (5-7 repeats) and 'short' (2-4 repeats), a significant excess of long alleles was observed in the patient group (P = 0.023, one-tailed), with an odds ratio of 2.30 (95% CI 1.07-4.93). We conclude that our findings support the hypothesis that alleles of the DRD4 exon III VNTR are susceptibility factors for heroin abuse.
The ryanodine receptor (RyR)/Ca2+ release channel is an essential component of excitation-contraction coupling in striated muscle cells. To study the function and regulation of the Ca2+ release channel, we tested the effect of caffeine on the full-length and carboxyl-terminal portion of skeletal muscle RyR expressed in a Chinese hamster ovary (CHO) cell line. Caffeine induced openings of the full length RyR channels in a concentration-dependent manner, but it had no effect on the carboxyl-terminal RyR channels. CHO cells expressing the carboxyl-terminal RyR proteins displayed spontaneous changes of intracellular [Ca2+]. Unlike the native RyR channels in muscle cells, which display localized Ca2+ release events (i.e., "Ca2+ sparks" in cardiac muscle and "local release events" in skeletal muscle), CHO cells expressing the full length RyR proteins did not exhibit detectable spontaneous or caffeine-induced local Ca2+ release events. Our data suggest that the binding site for caffeine is likely to reside within the amino-terminal portion of RyR, and the localized Ca2+ release events observed in muscle cells may involve gating of a group of Ca2+ release channels and/or interaction of RyR with muscle-specific proteins.
PURPOSE: Many systemic and topical therapeutic agents such as growth hormone, platelet-derived growth factor (PDGF), fibroblast growth factor (FGF), epidermal growth factor (EGF), and insulin-like growth factor (IGF) have been used as vulnerary agents. However, the role of nitric oxide (NO) as a wound-healing stimulant has been received with mixed reviews. NO is a potent vasodilator that is thought to be an endothelium-dependent relaxing factor, and a regulator of blood pressure and regional blood flow. It affects vascular smooth muscle proliferation and inhibits platelet aggregation and leukocyte adhesion. Therefore we compared the effects of several topical substances that have similar or reverse properties. METHODS: Using the excisional rat wound model, we evaluated the topical effects of Dermaide Aloe (D-Aloe, Dermaide Research Corp, Palos Heights, IL), nitroglycerin, Aquaphor (Beuersdorf, Inc., Norwalk, CT) alone, with D-Aloe with nitroglycerin, 2%, and L-NAME (NO inhibitor) with Aquaphor, and L-NAME with Aquaphor and D-Aloe for a 21-day period. All wounds were measured by planimetry at 1, 7, 10, 13, 16, 18, and 21 days. RESULTS: At day 1, all wounds had an average wound size of 2.27 cm2 (SD +/- 0.372) with no significant difference in wound size among the groups. Topically applied D-Aloe appeared to promote wound healing faster than the remaining other topicals (p < .05, Student-Newman-Keuls and Dunn's Method) over the study period. However, topicals combined with D-Aloe, the vehicle Aquaphor, and L-NAME improved the wound healing process when compared with nitroglycerin alone (p < .05). CONCLUSIONS: D-Aloe appears to have a wound-healing advancement factor that can reverse the effects of petrolatum- and nitroglycerin-based products as observed in the remaining groups when compared with nitroglycerin alone. It appears that D-Aloe's effect of preventing dermal ischemia by reversing the effects of thromboxane synthetase (TxA2) may act synergistically with NO or could be an oxygen radical scavenger.
BACKGROUND: The present paper addressed the issue of whether pretreatment with intravenous (i.v.) chemotherapy affects response rate or survival in patients receiving hepatic artery chemotherapy (HAC). METHODS: Case note reviews of 164 patients treated in a teaching hospital from June 1990 to July 1996 were carried out. RESULTS: The response rate and carcino-embryonic antigen (CEA) fall in the two groups was almost identical. There was a nonsignificant survival advantage in the non-pretreatment group. CONCLUSIONS: Previous administration of i.v. chemotherapy did not affect the CEA response of patients receiving HAC.
Coadministration of antagonists of N-methyl-D-aspartate (NMDA) receptor and opioids has been shown to prevent development of opiate tolerance in animal and clinical studies, but its cellular and molecular mechanisms are not understood. In this study, the effect of NMDA on delta-opioid receptor (DOR)-mediated signal transduction was investigated in neuroblastoma x glioma NG108-15 cells that functionally express both DOR and NMDA receptors. Acute incubation of NG108-15 cells with NMDA, a specific agonist of NMDA receptor, significantly attenuated the ability of DOR agonist [D-Pen2, D-Pen5]-enkephalin (DPDPE) to inhibit forskolin-stimulated cAMP production. The attenuation caused by NMDA was dose-dependent, and the EC50 of DPDPE increased 100-fold (from 4.6 nM to 500 nM) after NMDA treatment. The NMDA effect on responsiveness of delta-opioid receptors to DPDPE could be blocked by ketamine, a NMDA receptor-specific antagonist. This NMDA attenuation effect on DOR activity was also observed in neuronal primary cell cultures from fetal mouse brain but not in the Chinese hamster ovary cell line stably transfected with DOR alone. Interestingly, NMDA pretreatment reduced the cellular response to epinephrine but not to that of prostaglandin E1 in NG108-15 cells, which suggests differential modulation of NMDA on different G protein-coupled receptors. Pretreatment of NG108-15 cells with ketamine along with DPDPE greatly attenuated DPDPE-induced acute desensitization of DOR. Furthermore, the specific inhibitors of protein kinase C, either chelerythrine chloride or Go 6979, effectively blocked the NMDA effect, which indicates the involvement of protein kinase C in the process. In conclusion, the activation of NMDA receptors can attenuate acute responsiveness of DOR in neuronal cells, whereas its blockage leads to reduction of DOR desensitization. These results have thus provided an insight into cross-talk between NMDA and opioid signal transduction.
Matrix Gla protein (MGP) is a vitamin K-dependent extracellular matrix protein with a wide tissue distribution. The current study was designed to investigate the possible regulation of MGP by exogenous transforming growth factor-beta (TGF-beta) during lung development. Using reverse transcription coupled competitive polymerase chain reaction methodology, we determined that exogenous TGF-beta 1 increases MGP mRNA levels in embryonic mouse lung culture in a concentration-dependent manner. MGP mRNA levels were elevated by 5.0-fold at 50 ng/ml TGF-beta 1 in E11 embryonic mouse lungs explanted for 4 days in serumless culture. MGP mRNA stimulation by TGF-beta 1 was a time-dependent event: MGP message increment was initially detected after 1 day in culture, and MGP mRNA levels continued to increase up to 4 days in the presence of TGF-beta 1. In addition, embryonic lungs in serumless medium without exogenously added TGF-beta 1 showed an increase, although to a lesser extent, in pulmonary MGP mRNA level during culture, indicating that MGP expression is also developmentally regulated. The present results indicate that MGP gene expression can be stimulated by exogenous TGF-beta 1 during early embryonic mouse lung branching morphogenesis in culture.