Induction of caspase activation and cleavage of the viral nucleocapsid protein in different cell types during Crimean-Congo hemorrhagic fever virus infection

Regulation of apoptosis during infection has been observed for several viral pathogens. Programmed cell death and regulation of apoptosis in response to a viral infection are important factors for host or virus survival. It is not known whether Crimean-Congo hemorrhagic fever virus (CCHFV) infection regulates the apoptosis process in vitro. This study for the first time suggests that CCHFV induces apoptosis, which may be dependent on caspase-3 activation. This study also shows that the coding sequence of the S segment of CCHFV contains a proteolytic cleavage site, DEVD, which is conserved in all CCHFV strains. By using different recombinant expression systems and site-directed mutagenesis, we demonstrated that this motif is subject to caspase cleavage. We also demonstrate that CCHFV nucleocapsid protein (NP) is cleaved into a 30-kDa fragment at the same time as caspase activity is induced during infection. Using caspase inhibitors and cells lacking caspase-3, we clearly demonstrate that the cleavage of NP is caspase-3-dependent. We also show that the inhibition of apoptosis induced progeny viral titers of ～80-90%. Thus, caspase-3-dependent cleavage of NP may represent a host defense mechanism against lytic CCHFV infection. Taken together, these data suggest that the most abundant protein of CCHFV, which has several essential functions such as protection of viral RNA and participation in various processes in the replication cycle, can be subjected to cleavage by host cell caspases.     

Molecular mimicry of human endothelial cell antigen by autoantibodies to nonstructural protein 1 of dengue virus

The pathogenesis of dengue hemorrhagic fever and dengue shock syndrome (DHF/DSS), both serious complications of dengue virus (DV) infection, remains unclear. In this study, we found that anti-DV NS1 (nonstructural protein 1) polyclonal antibodies cross-reacted with human umbilical vein endothelial cells (HUVECs). We further identified a complex-specific mAb, DB16-1, which could recognize DV NS1 and cross-react with HUVECs and human blood vessels. The target protein of DB16-1 was further purified by immunoaffinity chromatography. LC-MS/MS analysis and co-immunoprecipitation revealed that the target protein of DB16-1 was human LYRIC (lysine-rich CEACAM1 co-isolated). Our newly generated anti-LYRIC mAbs bound to HUVECs in a pattern similar to that of DB16-1. The B-cell epitope of DB16-1 displayed a consensus motif, Lys-X-Trp-Gly (KXWG), which corresponded to amino acid residues 116-119 of DV NS1 and mimicked amino acid residues 334-337 in LYRIC. Moreover, the binding activity of DB16-1 in NS1 of DV-2 and in LYRIC disappeared after the KXWG epitope was deleted in each. In conclusion, DB16-1 targeted the same epitope in DV NS1 and LYRIC protein on human endothelial cells, suggesting that it might play a role in the pathogenesis of DHF/DSS. Future studies on the role of the anti-NS1 antibody in causing vascular permeability will undoubtedly be performed on sera collected from individuals before, during, and after the endothelial cell malfunction phase of a dengue illness.     

Thermal physiology of laboratory mice: Defining thermoneutrality

In terms of total number of publications, the laboratory mouse (Mus musculus) has emerged as the most popular test subject in biomedical research. Mice are used as models to study obesity, diabetes, CNS diseases and variety of other pathologies. Mice are classified as homeotherms and regulate their core temperature over a relatively wide range of ambient temperatures. However, researchers find that the thermoregulatory system of mice is easily affected by drugs, chemicals, and a variety of pathological conditions, effects that can be exacerbated by changes in ambient temperature. To this end, a thorough review of the thermal physiology of mice, including their sensitivity and regulatory limits to changes in ambient temperature is the primary focus of this review. Specifically, the zone of thermoneutrality for metabolic rate and how it corresponds to that for growth, reproduction, development, thermal comfort, and many other variables is covered. A key point of the review is to show that behavioral thermoregulation of mice is geared to minimize energy expenditure. Their zone of thermal comfort is essentially wedged between the thresholds to increase heat production and heat loss; however, this zone is above the recommended guidelines for animal vivariums. Future work is needed to better understand the behavioral and autonomic thermoregulatory responses of this most popular test species.     

Interleukin 1 reduces opioid binding in guinea pig brain

Interleukin 1 (IL1) is a macrophage-derived polypeptide which signals neurons in the preoptic-anterior hypothalamus to initiate fever and the acute-phase glycoprotein response. Recently, increases in cerebrospinal fluid and hypothalamic levels of β-endorphin have been reported during endotoxin (LPS)- and IL1-induced fevers, suggesting that this opioid may participate in the modulation of IL1 effects in the CNS. In this study, we investigated whether purified (human) IL1 influences the specific binding of three prototypic opioid agonists (2-D-alanine-5-L-methionineamide, DAME; (−)-ethylketocyclazocine, EKC; dihydromorphine, DHM) and one antagonist (naloxone) to opioid receptor-enriched membrane preparations in cerebral cortex, hypothalamus, midbrain, pons, medulla, and cerebellum of guinea pig brain. IL1 reduced the binding of these ligands to their receptors during a 30-min incubation. The extent of IL1 inhibition of a given ligand for its binding sites varied according to the brain region; within some regions, the extent of this inhibition also varied with the four ligands tested. But in cortex the effect of IL1 on the specific binding of DHM is dose-dependent. Similar results were obtained with crude homologous IL1. S. enteritidis endotoxin, suspended in pyrogen-free saline at concentrations from 4 to 36 μg/ml, did not inhibit the binding of these opioid ligands to their receptors in any brain region. These results indicate that IL1 interacts with the opiate receptors in guinea pig brain. This interaction, moreover, is not limited to the hypothalamus alone, the primary site of the pyrogenic action of IL1, but also occurs in other brain regions.     

2 152例门诊发热患者情况分析

目的 分析2152例门诊发热患者情况,探讨发热患者年龄分布;发热疾病的比率;呼吸道传染病对发热患者门诊就诊率的影响以及就诊前后用药情况。方法2152例患者为2003年5月～2004年5月在大连市开发区医院成人发热门诊就诊的全部发热患者,统计发热患者的平均年龄及性别差异;计算门诊初步诊断各疾病占总发热患者的百分率;计算发热患者占总门诊量的百分率;分析呼吸道传染病对发热患者门诊就诊率的影响;统计发热患者来院前的用药情况及门诊用药情况。结果 成人发热门诊患者,年龄15～90岁,平均年龄26．37岁;其中男性1150例女性1002例,各占总发热患者的53．44％、46．56oA;发热患者占同期医院总门诊量的0．82％,在“SARS”严重的2003年5、6月和禽流感流行的2004年1月发热患者分别占各月总门诊量的3．71％、2．51％、1．31％;发热患者中首诊为急性上呼吸道感染1379例,占总发热患者的64．03％,其中扁桃体炎397例,占急性上呼吸道感染的28．79％;其他发热患者首诊依次为急性胃肠炎157例,肺炎148例,支气管炎40例,肺结核20例,泌尿系感染8例,急性阑尾炎8例,妇科炎症5例,其他皮肤感染、疥、痛等明确原因的18例（其中流脑1例）,首诊未明确发热原因的18例。发热患者中WBC升高的348例,占总发热患者的16．17％;来院就诊前应用抗炎退热药者528例,占总发热患者的24．54％;在院静点抗菌药物者l052例,占总发热患者的48．88％,其中应用喹诺酮者529例,占总用药患者的50．28％,应用大环内脂类的511例（主要为阿奇霉素）,占总用药患者的48．57％,应用氨基糖苷类的8例,应用β-内酰胺类的4例;在用药者中,联合其他中成药或抗病毒药者122例,占总用药患者的11．59％。结论 成人门诊发热患者以青壮年为主,男性略多于女性;呼吸道传染病的流行大大增强了人们对发热的重视程度,到医院发热门诊就诊人数明显增多;在门诊就诊的发热患者中以急性上呼吸道感染为主,其他依次为急性胃肠炎、肺炎、支气管炎、肺结核等;门诊发热患者并非以细菌感染为主;患者就诊前、后不合理应用抗菌药物情况相当严重。     

Construction of an infectious cDNA clone of Tembusu virus isolated from breeder Peking ducks

正Dear Editor,Since April 2010,an outbreak of a new disease has elicited symptoms of high fever,loss of appetite,and reduction in egg production in layer ducks in eastern China;this phenomenon has now spread throughout China(Cao et al.,2011;Su et al.,2011).The causative agent of the disease was identified as Tembusu virus(TMUV),which was classified into the genus Flavivirus,     

Heat stress increases the rate of tolerance development to lipopolysaccharide in rats

We have investigated the effect of prior exposure of rats to either a hot environment (40°C, 2.6 kPa water vapor pressure) or a control environment (24°C, 1.9 kPa water vapor pressure) on the development of tolerance to lipopolysaccharide. Sixty minutes of heat exposure significantly raised the abdominal temperature of the heat-stressed rats above that of the control animals for a period of 80 min (P<0.001). However, neither the magnitude nor the time-course of the febrile response of the heat-stressed rats was significantly different from that of the control animals when an initial injection of lipopolysaccharide was given 24 h after temperature exposure (P>0.05). Nevertheless, heat exposure did increase the rate of tolerance development to successive lipopolysaccharide injections, spaced 3 days apart. Heat-stressed animals were tolerant by the second injection of lipopolysaccharide, whilst the control animals were tolerant only by the third injection of lipopolysaccharide. Therefore, heat stress increases the rate of tolerance development to lipopolysaccharide without affecting the response of heat-stressed animals to their first lipopolysaccharide injection.     

A new method based on entropy theory for genomic sequence analysis

We have refined entropy theory to explore the meaning of the increasing sequence data on nucleic acids and proteins more conveniently. The concept of selection constraint was not introduced, only the analyzed sequences themselves were considered. The refined theory serves as a basis for deriving a method to analyze non-coding regions (NCRs) as well as coding regions. Positions with maximal entropy might play the most important role in genome functions as opposed to positions with minimal entropy. This method was tested in the well-characterized coding regions of 12 strains of Classical Swine Fever Virus (CSFV) and non-coding regions of 20 strains of CSFV. It is suitable to analyze nucleic acid sequences of a complete genome and to detect sensitive positions for mutagenesis. As such, the method serves to formulate the basis for elucidating the functional mechanism.     

Effect of a mu-opioid receptor-selective antagonist on interleukin-6 fever

The endogenous opioid system has been found to be involved in fever caused by pyrogens. Recent work in our laboratory has demonstrated that the mu-opioid receptor is involved in interleukin-1beta (IL-1beta)- and in lipopolysaccharide (LPS)-induced fevers. In the present study, we have investigated the role of the mu-opioid receptor in the preoptic anterior hypothalamus (POAH) in fever induced by interleukin-6 (IL-6). Following stereotaxic implantation of a guide cannula into the POAH for microinjection, radio transmitters to monitor body temperature (Tb) continuously were inserted intraperitoneally. Adult male Sprague-Dawley rats were microinjected with 0.5 microg of the selective mu-opioid receptor antagonist, cyclic D-phe-Cys-Try-D-Trp-Arg-Thr-Pen-Thr-NH2 (CTAP), into the POAH. Thirty min later, IL-6 (100 ng) was injected into the POAH. CTAP significantly blocked the IL-6 fever. CTAP alone had no effect on Tb during the 390-min recording period. These data indicate that mu-opioid receptors within the POAH mediate IL-6 fever and add to the increasing evidence that the opioid system is involved in the pathogenesis of fever in rats.