Navigating Transcriptional Coregulator Ensembles to Establish Genetic Networks: A GATA Factor Perspective.

Complex developmental programs require orchestration of intrinsic and extrinsic signals to control cell proliferation, differentiation, and survival. Master regulatory transcription factors are vital components of the machinery that transduce these stimuli into cellular responses. This is exemplified by the GATA family of transcription factors that establish cell type-specific genetic networks and control the development and homeostasis of systems including blood, vascular, adipose, and cardiac. Dysregulated GATA factor activity/expression underlies anemia, immunodeficiency, myelodysplastic syndrome, and leukemia. Parameters governing the capacity of a GATA factor expressed in multiple cell types to generate cell type-specific transcriptomes include selective coregulator usage and target gene-specific chromatin states. As knowledge of GATA-1 mechanisms in erythroid cells constitutes a solid foundation, we will focus predominantly on GATA-1, while highlighting principles that can be extrapolated to other master regulators. GATA-1 interacts with ubiquitous and lineage-restricted transcription factors, chromatin modifying/remodeling enzymes, and other coregulators to activate or repress transcription and to maintain preexisting transcriptional states. Major unresolved issues include: how does a GATA factor selectively utilize diverse coregulators; do distinct epigenetic landscapes and nuclear microenvironments of target genes dictate coregulator requirements; and do gene cohorts controlled by a common coregulator ensemble function in common pathways. This review will consider these issues in the context of GATA factor-regulated hematopoiesis and from a broader perspective.

Effect of magnetic field on positron lifetimes of Fe, Co and Ni.

Positron lifetime spectra of Fe, Co and Ni were measured under magnetic field using a (22)Na source. Very small but distinguishable difference of positron lifetime upon magnetic field reversal was observed suggesting the existence of two bulk lifetimes associated with majority and minority spin electrons. Using two spin-dependent Fe bulk lifetimes, the difference Doppler broadening of annihilation radiation spectra between majority and minority spin electrons were also examined. Agreement between experiment and theory indicates that spin-polarized positron annihilation spectroscopy may have potential in investigation of spin-aligned electron momentum distribution.

[Coronary artery bypass grafting for unstable angina pectoris with aortitis syndrome; report of a case].

A 78-year-old male with aortitis syndrome was referred to our hospital for the treatment of unstable angina pectoris with ischemic mitral regurgitation, which was diagnosed by transthoracic echocardiography and coronary artery angiography. Computed tomography showed segmental wall thickness of thoracic and abdominal aorta He underwent an emergent coronary artery bypass grafting. The postoperative course was uneventful without any neurological complications. Postoperative echocardiogram and coronary artery angiography showed good mitral valve function and all patent bypass grafts. He was discharged 33 days after surgery. At 26 months after surgery, he is well without limitation of daily activities and any evidence of myocardial ischemia.

[Surgical cryoablation and left ventriculoplasty for electrical storm after acute myocardial infarction].

A 65-year-old man was referred to our hospital to treat recent anterior myocardial infarction. Coronary artery angiography showed acute occlusion of left anterior descending coronary artery (LAD) and chronic occlusion of right coronary artery. After emergent percutaneous coronary intervention for LAD, drug-refractory electrical storm necessitating frequent electrical defibrillating cardioversion occurred. This patient successfully underwent surgical cryoablation, left ventriculoplasty and coronary revascularization. At 2 years and 10th month after the operation, he is well without limitation of daily activities and any evidence of myocardial ischemia and ventricular tachycardia.

[Surgery for multiple aortic aneurysms with unstable angina pectoris; report of a case].

A 77-year-old female with unstable angina pectoris was referred to our hospital for further evaluation of multiple aortic aneurysms. Computed tomography showed descending thoracic (65 mm), thoracoabdominal (40 mm) and infra-renal abdominal aneurysm (50 mm). Initially, this patient underwent off pump coronary revascularization. On 11 days after initial surgery, descending thoracic aneurysm ruptured, followed by emergent descending thoracic and thoraco-abdominal aneurysm repair. Two months later from this aortic repair, this patient successfully underwent abdominal aortic aneurysm repair. At 3 years and 7th month after the last operation, she is well without limitation of daily activities and any evidence of myocardial ischemia.

[On pump beating heart mitral valve repair without aortic cross-clamp for low ejection fraction ischemic mitral regurgitation after coronary artery bypass grafting; report of a case].

A 59-year-old male with congestive heart failure caused by impaired left ventricular function after coronary artery bypass grafting (CABG) was referred to our hospital, and massive ischemic mitral regurgitation was detected by echocardiography. This patient underwent on-pump beating-heart mitral valve repair without aortic cross-clamp successfully through right thoracotomy. Postoperative echocardiography revealed no mitral regurgitation. The patient recovered uneventfully and was discharged on the 17th postoperative day. At 6th month after the operation, he is well without mitral regurgitation.

Impact of different husbandry conditions on contact and airborne transmission of H5N1 highly pathogenic avian influenza virus to chickens.

Typically highly pathogenic avian influenza (HPAI) viruses spread very rapidly among chickens within sheds. However, the spread was slower than expected for the initial 10 days of the index farm in Japan during 2004. This slow spread, as well as the lack of gross lesions, clinical signs, or high mortality, hindered the field veterinarian from reporting a suspected HPAI outbreak to the veterinary office. To understand the field conditions for the slow virus spread, we examined contact and airborne transmission of the H5N1 virus to chickens in a negative-pressure isolator using various numbers of infected chickens and separate compartments. We found that the contact transmission did occur inefficiently when one or two chickens were infected, whereas the transmission was efficient when four chickens were infected. Airborne transmission of the HPAI virus was also dependent on the number of infected chickens and was less efficient than contact transmission. These data together with field observations suggested that number of infected chickens, chicken house types, and amount of environmental contamination might affect the virus transmission efficiency to chickens.

Distribution of viral antigens and development of lesions in chicken embryos inoculated with nipah virus.

An isolate of Nipah virus was injected into fertile eggs via the allantoic cavity or yolk sac. Allantoic inoculation resulted in considerable pathological variation and only partial mortality. Dead embryos showed severe necrosis in the brain and congestion in the kidney and the subcutis of limbs. In contrast, yolk sac inoculation led to uniform infection and mortality, the dead embryos exhibiting the same lesions as those described above but without the subcutaneous congestion. Histological lesions in dead embryos inoculated by either route were similar and particularly severe in the central nervous system. Viral antigens were detected mainly in the vasculature and neurons. The results indicated that Nipah virus is highly pathogenic to chicken embryos, and that the route of inoculation is an important determinant of the course of disease. The findings also suggested that yolk sac inoculation can be used for viral titration, and that the chicken embryo represents a useful model for studying the vascular and neuronal tropisms of Nipah virus.

Pathology of fatal highly pathogenic H5N1 avian influenza virus infection in large-billed crows (Corvus macrorhynchos) during the 2004 outbreak in Japan.

Highly pathogenic H5N1 avian influenza viruses were isolated in 9 large-billed crows that died in Kyoto and Osaka prefectures in Japan from March to April in 2004. We studied 3 of the 9 crows using standard histologic methods, immunohistochemistry, and virus isolation. The most prominent lesions were gross patchy areas of reddish discoloration in the pancreas. The consistent histologic lesions included severe multifocal necrotizing pancreatitis, focal degeneration and necrosis of neuron and glial cells in the central nervous system, and focal degeneration of cardiac myocytes. All of these tissues contained immunohistochemically positive influenza viral antigens. The virus was isolated from the brain, lung, heart, liver, spleen, and kidney of the crows examined. Thus we concluded that highly pathogenic avian influenza virus was associated with clinical disease, severe pathologic changes, and death in the 3 crows.

Monoclonal antibody-based immunohistochemical diagnosis of Malaysian Nipah virus infection in pigs.

Formalin-fixed, paraffin wax-embedded tissues of three Malaysian farm pigs naturally infected with Nipah virus were used to investigate the value of anti-Nipah virus mouse monoclonal antibodies (Mabs) and rabbit polyclonal antibody for immunohistochemical diagnosis. Mabs 11F6 and 12A5 gave intense immunolabelling in lung tissue that had been fixed in 10% neutral buffered formalin for about 4 years, whereas the reactivity of Mabs 13A5 and 18C4 and polyclonal antibody was reduced significantly by long-term formalin fixation. Immunohistochemical examination of Malaysian farm pig samples with Mab 11F6 confirmed the affinity of Nipah virus for respiratory epithelium, renal glomerular and tubular epithelium, meningeal arachnoidal cells, and systemic vascular endothelium and smooth muscle. In addition, Nipah virus antigens were identified in laryngeal epithelial cells, Schwann cells of peripheral nerve fascicles in the spleen, and endothelial cells in the atrioventricular valve. The study demonstrated the value of Mabs 11F6 and 12A5 for the immunohistochemical diagnosis of Nipah virus infection in pigs.

Complete, long-lasting protection against lethal infectious bursal disease virus challenge by a single vaccination with an avian herpesvirus vector expressing VP2 antigens.

Marek's disease herpesvirus is a vaccine vector of great promise for chickens; however, complete protection against foreign infectious diseases has not been achieved. In this study, two herpesvirus of turkey recombinants (rHVTs) expressing large amounts of infectious bursal disease virus (IBDV) VP2 antigen under the control of a human cytomegalovirus (CMV) promoter or CMV/beta-actin chimera promoter (Pec promoter) (rHVT-cmvVP2 and rHVT-pecVP2) were constructed. rHVT-pecVP2, which expressed the VP2 antigen approximately four times more than did rHVT-cmvVP2 in vitro, induced complete protection against a lethal IBDV challenge in chickens, whereas rHVT-cmvVP2 induced 58% protection. All of the chickens vaccinated with rHVT-pecVP2 had a protective level of antibodies to the VP2 antigen at the time of challenge, whereas only 42 and 67% of chickens vaccinated with rHVT-cmvVP2 or the conventional live IBDV vaccine, respectively, had the antibodies. The antibody level of chickens vaccinated with rHVT-pecVP2 increased for 16 weeks, and the peak antibody level persisted throughout the experiment. The serum antibody titer at 30 weeks of age was about 20 or 65 times higher than that of chickens vaccinated with rHVT-cmvVP2 or the conventional live vaccine, respectively. rHVT-pecVP2, isolated consistently for 30 weeks from the vaccinated chickens, expressed the VP2 antigen after cultivation, and neither nucleotide mutations nor deletion in the VP2 gene was found. These results demonstrate that the amount of VP2 antigen expressed in the HVT vector was correlated with the vaccine efficacy against lethal IBDV challenge, and complete protective immunity that is likely to persist for the life of the chickens was induced.

Epizootic outbreaks of gizzard erosion associated with adenovirus infection in chickens.

Two outbreaks of gizzard erosion in slaughtered broiler chickens in Japan were examined pathologically and microbiologically. The prevalences of such lesions were 9%-11% and 4%-50% in the affected flocks. Affected chickens had no clinical signs. Group I fowl adenovirus (FAV) serotype 1 was isolated from gizzard lesions. Histologically, gizzard mucosa were necrotic. Intranuclear inclusion bodies were seen in the enlarged nuclei of degenerating epithelial cells of the gizzard. The keratinoid layer in the erosion was edematous and desquamated and contained degenerative cells. Moderate to marked inflammatory cell infiltration was observed in the lamina propria and perivascular connective tissue in the submucosa and muscle layer. Immunohistochemical staining showed evidence of FAV antigens in the intranuclear inclusion bodies within degenerating epithelial cells. Ultrastructurally, numerous viral particles were demonstrated in the inclusions.

Identification of a genetic determinant of pathogenicity in chicken anaemia virus.

The molecular basis of pathogenicity of the chicken anaemia virus (CAV) needs to be clarified in order to develop a safe, live virus vaccine. In this study, several high- and low-pathogenic infectious DNA clones were obtained from field virus samples after 12 or 38 passages in MDCC-MSB1 cells. The high-pathogenic clones induced a low haematocrit, low weight gain and high mortality. Nucleotide sequence analyses identified one amino acid, at residue 394 of the VP1 capsid protein, as a major determinant of pathogenicity. To determine the role of this amino acid in pathogenicity, chimeric infectious DNA clones and point-mutated clones were used for chicken pathogenicity tests. These analyses clearly demonstrated that residue 394 of VP1 was crucial for the pathogenicity of CAV; all of the cloned viruses with glutamine at this position were highly pathogenic, whereas those with histidine had low pathogenicity. Low-pathogenic CAV, based on an infectious DNA clone, is a candidate for a genetically homogeneous and stable CAV live vaccine.

Immunohistochemical characterization of calf pneumonia produced by the combined endobronchial administration of bovine herpesvirus 1 and Pasteurella haemolytica.

Ten calves ("group 4") were inoculated endobronchially with Pasteurella haemolytica 4 days after inoculation with bovine herpesvirus 1 (BHV-1). Four calves (group 3) were similarly inoculated with P. haemolytica alone, and three (group 2) with BHV-1 alone. All group 4 animals showed severe respiratory signs and had bilateral lobar pneumonia; one died 6 days after inoculation with P. haemolytica. Two types of pneumonic lesion were observed. One was characterized by interlobular and interstitial lymphatic thrombosis, fibrinous pleuritis and coagulative necrosis, and the other by necrotizing bronchiolitis with intranuclear inclusion bodies. The former type of lesion was associated with the presence of P. haemolytica antigen and the latter with the presence of BHV-1 antigen. The weight of infection of BHV-1 and P. haemolytica in bronchoalveolar (BAL) fluid was clearly reflected in the immunohistochemical demonstration of the corresponding antigens in BAL fluid cells. Group 4 calves differed from the calves of groups 1-3 in showing 10-1530 times more endotoxin in BAL fluid. These findings suggested that BHV-1 infection partly destroyed the clearance mechanisms of the respiratory tract epithelium and exacerbated the subsequent P. haemolytica infection.

Brain lesions and transmission of experimental equine herpesvirus type 9 in pigs.

We demonstrated that pigs are susceptible to acute infection by equine herpesvirus type 9 (EHV-9). Six 8-week-old SPF pigs were inoculated intranasally and four were inoculated orally with different doses of EHV-9, and observed for 6 days. Although neurological signs did not develop in any of the infected pigs, the six intranasally infected pigs and one of the orally infected pigs developed lesions of encephalitis consisting of neuronal necrosis, neuronophagia, and intranuclear inclusion bodies, distributed mainly in the rhinencephalon. EHV-9 antigen was localized in the necrotic neuronal cells and was closely associated with the presence of inclusion bodies. These findings clearly demonstrate that pigs are fully susceptible to EHV-9 infection following intranasal inoculation (but less so following oral inoculation), and that EHV-9 in pigs has a highly neurotropic nature.

Pneumonia induced byEndobronchial inoculation of calves with bovine herpesvirus 1.

Each of six calves inoculated endobronchially with bovine herpesvirus 1 (BHV-1) by means of a bronchoscope developed viral pneumonia. Gross and histopathological lesions were mainly localized to the right diaphragmatic lobe (middle to caudal region) of the lung and were closely associated with the site of the deposition of the inoculum. The lesions were characterized by intranuclear inclusion bodies associated with focal necrosis of the epithelium in the lower respiratory tract. BHV-1 antigen and BHV particles were detected in the degenerating bronchial, bronchiolar and alveolar epithelial cells. After infection, the total cell count in the broncho-alveolar lavage (BAL) fluid increased. In addition, BHV-1 antigen and virus were detected in the desquamated cells and macrophages of BAL fluid from the right diaphragmatic lobe, but not from the left diaphragmatic lobe. It is concluded that examination of BAL fluid is valuable for immunohistopathological and virological confirmation of BHV-1 infection.

Long-term culture of glutamine synthetase-transfected HepG2 cells in circulatory flow bioreactor for development of a bioartificial liver.

Glutamine synthetase (GS) is involved in an accessory pathway of ammonia removal in mammals. To develop a bioartificial liver with a human cell line, GS gene was transfected into HepG2 cells, which had no ammonia removal activity. After culturing in the presence of methionine sulfoximine (MSX), a GS inhibitor, we obtained a MSX-resistant HepG2 subline (GS-HepG2), which had amplified GS gene; ammonia removal activity was estimated to be 1/7 of that of rat primary culture hepatocytes. The cells were cultured in a circulatory flow bioreactor for 109 days, while they multiplied from 5 x 10(7) to 4 x 10(9) cells. Three days after inoculation, the ammonia level of the culture medium was lowered to a level maintained thereafter, suggesting that using recombinant cell lines for bioartificial livers enables long-term repeated treatment for hepatic failure patient. Judging from the rate of decrease in the amount of the added ammonia, the ammonia removal capability of 4 x 10(9) GS-HepG2 cells was almost equivalent to 5 x 10(8) porcine hepatocytes inoculated into the circulatory flow bioreactor. Apart from their ammonia removal activity, GS-HepG2 cells eliminated human tumor necrosis factor-alpha (TNF-alpha). Cytokine removal therefore promises to be another useful property of bioreactor cells.

Dual-viral vector approach induced strong and long-lasting protective immunity against very virulent infectious bursal disease virus.

To induce strong protective immunity against very virulent infectious bursal disease virus (vvIBDV) in chickens, two viral vector systems, Marek's disease and Fowlpox viruses expressing the vvIBDV host-protective antigen VP2 (rMDV, rFPV), were used. Most of chickens vaccinated with the rFPV or rMDV alone, or vaccinated simultaneously with both at their hatch (rMDV-rFPV(1d)), were protected against developing clinical signs and mortality; however, only zero to 14% of the chickens were protected against gross lesions. In contrast, gross lesions were protected in 67% of chickens vaccinated primarily with the rMDV followed by boosting with the rFPV 2 weeks later (rMDV-rFPV(14d)). Protection against the severe histopathological lesions of rFPV, rMDV, rMDV-rFPV(1d), and rMDV-rFPV(14d) vaccine groups were 33, 42, 53, and 73%, respectively. Geometric mean antibody titers to VP2 of chickens vaccinated with the rFPV, rMDV, rMDV-rFPV(1d), and rMDV-rFPV(14d) before the challenge were 110, 202, 254, and 611, respectively. Persistent infection of the rMDV in chickens after the booster vaccination with rFPV was suggested by detection of the rMDV genes from peripheral blood lymphocyte DNA at 28 weeks of age. These results indicate that the dual-viral vector approach is useful for quickly and safely inducing strong and long-lasting protective immunity against vvIBDV in chickens.

Immunohistochemical detection of hog cholera virus antigen in paraffin wax-embedded tissues from naturally infected pigs.

In 17 pigs submitted for diagnosis in 1980, hog cholera was confirmed by viral isolation, by a direct immunofluorescent antibody test for viral antigen, and by the presence of characteristic histopathological lesions. In the present study, hog cholera viral antigen was demonstrated in these pigs by immunohistochemical examination of formalin-fixed paraffin wax-embedded tissues that had been stored for 18 years. Viral antigen was detected in crypt epithelial cells of the tonsil, collecting tubular epithelial cells of the kidney, bronchial and bronchiolar mucosal and gland cells of the lung, and pancreatic epithelial cells.

Protection of chickens against very virulent infectious bursal disease virus (IBDV) and Marek's disease virus (MDV) with a recombinant MDV expressing IBDV VP2.

To develop a herpes virus vaccine that can induce immunity for an extended period, a recombinant Marek's disease (MD) virus (MDV) CVI-988 strain expressing infectious bursal disease virus (IBDV) host-protective antigen VP2 at the US2 site (rMDV) was developed under the control of an SV40 early promoter. Chickens vaccinated with the rMDV showed no clinical signs and no mortality and 55% of the chickens were considered protected histopathologically after challenge with very virulent IBDV (vvIBDV), whereas all of the chickens vaccinated with the conventional IBDV vaccine showed no clinical signs and were protected. Chickens vaccinated with the CVI-988 or chickens in the challenge control showed severe clinical signs and high mortality (70-75%) and none of them were protected. Also, the rMDV conferred full protection to chickens against vvMDV just as the CVI-988 strain did, whereas 90% of the challenge control chickens died of MD. Antibody levels against IBDV and MDV following the vaccination increased continuously for at least 10 weeks. No histopathological lesions in the rMDV-vaccinated chickens and no contact transmission of the rMDV to their penmates were confirmed. These results demonstrate that an effective and safe recombinant herpesvirus-based IBD vaccine could be constructed by expressing the VP2 antigen at the US2 site of the CVI-988 vaccine strain.