Lifespan Trajectories of White Matter Changes in Rhesus Monkeys.

Progress in neurodevelopmental brain research has been achieved through the use of animal models. Such models not only help understanding biological changes that govern brain development, maturation and aging, but are also essential for identifying possible mechanisms of neurodevelopmental and age-related chronic disorders, and to evaluate possible interventions with potential relevance to human disease. Genetic relationship of rhesus monkeys to humans makes those animals a great candidate for such models. With the typical lifespan of 25 years, they undergo cognitive maturation and aging that is similar to this observed in humans. Quantitative structural neuroimaging has been proposed as one of the candidate in vivo biomarkers for tracking white matter brain maturation and aging. While lifespan trajectories of white matter changes have been mapped in humans, such knowledge is not available for nonhuman primates. Here, we analyze and model lifespan trajectories of white matter microstructure using in vivo diffusion imaging in a sample of 44 rhesus monkeys. We report quantitative parameters (including slopes and peaks) of lifespan trajectories for 8 individual white matter tracts. We show different trajectories for cellular and extracellular microstructural imaging components that are associated with white matter maturation and aging, and discuss similarities and differences between those in humans and rhesus monkeys, the importance of our findings, and future directions for the field. Significance Statement: Quantitative structural neuroimaging has been proposed as one of the candidate in vivo biomarkers for tracking brain maturation and aging. While lifespan trajectories of structural white matter changes have been mapped in humans, such knowledge is not available for rhesus monkeys. We present here results of the analysis and modeling of the lifespan trajectories of white matter microstructure using in vivo diffusion imaging in a sample of 44 rhesus monkeys (age 4-27). We report and anatomically map lifespan changes related to cellular and extracellular microstructural components that are associated with white matter maturation and aging.

Right-sided hemiagenesis of the thyroid lobe and isthmus: A case report.

Unilateral or bilateral hypoplasia or agenesis of one or both thyroid lobes, with or without isthmic agenesis, is a rare developmental anomaly. Hemiagenesis of the left lobe is far commoner than of the right. Clinically, these patients may be euthyroid, hyperthyroid, or hypothyroid. Ultrasonography is usually able to diagnose this condition easily, as we demonstrate in this case report of a 37-year-old lady with an incidentally detected thyroid nodule who was found to have hemiagenesis of the right lobe and isthmus.

Need for an individualized and aggressive management of multinodular goiters of endemic zones by specially trained surgeons: experience in western Nepal.

BACKGROUND: The goals of the present study were to explore the presentation of multinodular goiter (MNG) and solitary thyroid nodules (STN) in the sub-Himalayan belt, including the risk of malignancy, and to evaluate whether specialized surgeon training in endocrine surgery has an effect on reducing complications. METHODS: This retrospective study (1998-2003) analyzed 624 patients with thyroid disorders seen in the thyroid clinic of a tertiary care hospital in western Nepal. The findings included 67.7% (n = 423: euthyroid, 297, toxic, 126) multinodular goiters (MNG) and 18.5% (n = 116) STN. Rest of patients of other thyroid disorders were excluded from the study. Ultrasonography and fine-needle aspiration cytology (FNAC) were the available diagnostic adjuncts. To evaluate the role of surgeon training, outcomes were compared between patients cared for by surgeons specially trained in endocrine surgery and those who were not. Prognostic markers indicated aggressiveness of cancers. RESULTS: Of the 539 MNG and STN patients in this series, 236 underwent operation. Of these, 25.7% (139/539) were toxic, and 11.31% had associated carcinoma. Aggressive cancers, like poorly differentiated (4.9%) and anaplastic types (18%), were more common than in series of patients from iodine-sufficient regions. Patients 40-55 years of age were more likely to have toxicity, and those > 60 years of age were more likely to have aggressive cancers. Postoperative complication rates were lower in the group treated by surgeons who had special training in endocrine surgery. CONCLUSIONS: There is a higher incidence of toxicity and malignancy in MNG in an endemic goiter zone. The limited diagnostic and therapeutic facilities in the region under study warrant a high degree of clinical suspicion and judgment, sound knowledge of thyroid physiology, thorough interpretation of hormone test results, and meticulous surgical techniques. The treatment must be individualized with consideration of humanitarian and socioeconomic factors, without compromising the quality of care and its long-term consequences. Aggressive management of malignancy and toxicity with total thyroidectomy is needed as primary therapy in many instances. However, subtotal excision is more useful in carefully selected cases with a small remnant. Specialized training in thyroid surgery appears to be valuable in reducing complications.

Mutational analysis of early region 4 of bovine adenovirus type 3.

The primary objective of characterizing bovine adenovirus type 3 (BAV3) in greater detail is to develop it as a vector for gene therapy and vaccination of humans and animals. A series of BAV3 early region 4 (E4) deletion-mutant viruses, containing deletions in individual E4 open reading frames (Orf) or combinations of Orfs, were generated by transfecting primary fetal bovine retinal cells with E4-modified genomic DNA. Each of these mutants was further analyzed for growth kinetics, viral DNA accumulation, and early-late protein synthesis. Mutant viruses carrying deletions in Orf1, Orf2, Orf3, or Orf4 showed growth characteristics similar to those of the E3-deleted BAV3 (BAV302). DNA accumulation and early/late protein synthesis were also indistinguishable from those of BAV302. However, mutant viruses carrying a deletion in Orf5, Orfs 1-3 (BAV429), or Orfs 3-5 (BAV430) were modestly compromised in their ability to grow in bovine cells and express early/late proteins. E4 mutants containing larger deletions, Orfs 1-3 (BAV429) and Orfs 3-5 (BAV430), were further tested in a cotton rat model. Both mutants replicated as efficiently as BAV3 or BAV302 in the lungs of cotton rats. BAV3-specific IgA and IgG responses were detected in serum and at the mucosal surfaces in cotton rats inoculated with mutant viruses. In vitro and in vivo characterization of these E4 mutants suggests that none of the individual E4 Orfs are essential for viral replication. Moreover, successful deletion of a 1.5-kb fragment in the BAV3 E4 region increased the available insertion capacity of replication-competent BAV3 vector (E3-E4 deleted) to approximately 4.5 kb and that of replication-defective BAV3 vector (E1a-E3-E4 deleted) to approximately 5.0 kb. This is extremely useful for the construction of BAV3 vectors that express multiple genes and/or regulatory elements for gene therapy and vaccination.

Juvenile rheumatoid arthritis with amyloid goiter: report of a case with review of the literature.

Clinically significant enlargement of the thyroid owing to amyloid deposition is a rare occurrence. A 23-yr-old female, a case of juvenile rheumatoid arthritis, developed rapidly increasing thyromegaly during the course of her illness with complaints of dyspnea and dysphagia. Thyroid function tests were within normal limits. Fine-needle aspiration cytology proved inconclusive. Total thyroidectomy was done for symptomatic relief with a preoperative clinical impression of malignancy. Histopathologic findings were consistent with amyloid goiter. The findings of this case are presented, to emphasize the difficulties in making a definite preoperative diagnosis, along with a brief review of the literature.

Recombinant bovine adenovirus type 3 expressing bovine viral diarrhea virus glycoprotein E2 induces an immune response in cotton rats.

Recombinant bovine adenovirus is being developed as a live vector for animal vaccination and for human gene therapy. In this study, two replication-competent bovine adenovirus 3 (BAV-3) recombinants (BAV331 and BAV338) expressing bovine viral diarrhea virus (BVDV) glycoprotein E2 in the early region 3 (E3) of BAV-3 were constructed. Recombinant BAV331 contains chemically synthesized E2 gene (nucleotides modified to remove internal cryptic splice sites) under the control of BAV-3 E3/major late promoter (MLP), while recombinant BAV338 contains original E2 gene under the control of human cytomegalovirus immediate early promoter. Since E2, a class I membrane glycoprotein, does not contain its own signal peptide sequence at the 5' end, the bovine herpesvirus 1 (BHV-1) glycoprotein D signal sequence was fused in frame to the E2 open reading frame (ORF) for proper processing of the E2 glycoprotein in both the recombinant viruses. Recombinant E2 protein expressed by BAV331 and BAV338 recombinant viruses was recognized by E2-specific monoclonal antibodies as a 53-kDa protein, which also formed dimer with an apparent molecular weight of 94 kDa. Insertion of an E2-expression cassette in the E3 region did not effect the replication of recombinant BAV-3s. Intranasal immunization of cotton rats with these recombinant viruses generated E2-specific IgA and IgG responses at the mucosal surfaces and in the serum. In summary, these results show that the pestivirus glycoprotein can be expressed efficiently by BAV-3. In addition, mucosal immunization with replication-competent recombinant bovine adenovirus 3 can induce a specific immune response against the expressed antigen.

Transcription map and expression of bovine herpesvirus-1 glycoprotein D in early region 4 of bovine adenovirus-3.

Early region 4 (E4) of bovine adenovirus type 3 (BAV-3) was analyzed by Northern blotting, RT-PCR analysis, cDNA sequencing, and S1 nuclease protection assays. The transcriptional map of the E4 region of BAV-3 has marked dissimilarities from those of mouse adenovirus-1, ovine adenovirus-287, and human adenovirus-2, for which the transcriptional maps have been constructed. The E4 region of BAV-3, located between 98.6 and 89.8 MU transcribes seven distinct classes of bovine adenovirus type 3 mRNA. The seven mRNA species formed by the removal of one to three introns share both the 3' end and a short 5' leader (25 nucleotides). The E4 mRNAs can encode at least five unique polypeptides, namely, 143R1, 69R, 143R2, 268R, and 219R. Isolation of a replication-competent recombinant "BAV404" containing 1.9-kb insertion [glycoprotein (gD) of bovine herpesvirus 1, under the control of a SV40 early promoter and poly(A)] in the region between E4 and the right ITR suggested that this region is nonessential for BAV-3 replication. Expression of gD by BAV404 recombinant virus was confirmed by immunoprecipitation with gD-specific monoclonal antibodies. Analysis of the kinetics of protein expression indicated that gD is expressed at both early and late times postinfection. These results suggest that: (a) E4 produces seven 5'-3' coterminal mRNAs and (b) the right terminal region of BAV-3 can be used for the expression of vaccine antigens.

Comparative evaluation of cell culture-adapted and chicken embryo-adapted fowl pox vaccine strains.

Two types of vaccines, chicken embryo adapted (VacCE) and cell culture adapted (VacCC), were tested for their efficacy to elicite the immune response in birds vaccinated at 2 and 8 wk of age. The cell-mediated immune response studied by blastogenesis assay showed that birds vaccinated at the second week of age by both VacCE and VacCC vaccines had significant increase in T-lymphocyte count at 21 days postvaccination (PV) and 7 days postchallenge (PC), whereas in birds vaccinated at 8 wk of age, a significant increase was seen at 21 days PV and 7 days PC with the VacCC vaccine. The rise in passive hemagglutination titers was observed up to 21 days PV and 7 days PC in birds vaccinated at 2 wk of age. However, only the birds vaccinated with VacCC at 8 wk of age showed rise in titers at days 21 PV and 7 PC. Birds were challenged 90 days PV by scarification on the thigh region, and the birds vaccinated with VacCC showed 90% and 70% protection when vaccinated at 2 and 8 wk, respectively. The birds vaccinated with VacCE showed only 60% and 20% protection at the corresponding levels, respectively.

Genetic organization and DNA sequence of early region 4 of bovine adenovirus type 3.

We have identified and sequenced 3614 nucleotides located at the extreme right-end of the bovine adenovirus type 3 (BAV3) genome from map units 89.5-100. Analysis of the sequence revealed an inverted terminal repeat (ITR) of 195 bp, and identified five open reading frames (ORFs) designated ORF1, ORF2, ORF3, ORF4 and ORF5. When compared with known E4 ORFs of other adenoviruses, ORFs 1, 2 and 4, which code for proteins of 143, 69 and 143 amino acids respectively, were found to be unique to BAV3. ORFs 3 and 5, which code for proteins of 268 and 219 amino acids respectively, showed partial homology to the E4 34 kDa protein of human adenovirus 2. Nucleotide sequence analysis also identified two potential TATA boxes upstream of ORF1 and a potential polyadenylation signal downstream of ORF5 suggesting that E4 transcripts may be 3' co-terminal.

Construction and characterization of E3-deleted bovine adenovirus type 3 expressing full-length and truncated form of bovine herpesvirus type 1 glycoprotein gD.

Using the homologous recombination machinery of E. coli, a 1.245-kb deletion was introduced in the E3 region of bovine adenovirus 3 (BAV3) genomic DNA cloned in a plasmid. Transfection of the restriction enzyme-excised, linear E3-deleted BAV3 genomic DNA into primary fetal bovine retina cells produced infectious virus (BAV3. E3d), suggesting that all the E3-specific open reading frames are nonessential for virus replication in vitro. Using a similar approach, we constructed replication-competent (BAV3.E3gD and BAV3. E3gDt) BAV3 recombinant expressing full-length (gD) or truncated (gDt) glycoprotein of bovine herpes virus 1. Recombinant gD and gDt proteins expressed by BAV3.E3gD and BAV3.E3gDt, respectively, were recognized by gD-specific monoclonal antibodies directed against conformational epitopes, suggesting that antigenicity of recombinant gD and gDt was similar to that of the native gD expressed in bovine herpes virus 1-infected cells. Intranasal immunization of cotton rats induced strong gD- and BAV3-specific IgA and IgG immune responses. These results suggest that replication-competent bovine adenovirus 3-based vectors have potential for the delivery of vaccine antigens to the mucosal surfaces of animals.

Characterization of bovine adenovirus type 3 early region 2B.

We have determined the nucleotide sequence of a 6999 base pair region of bovine adenovirus-3 covering map units 9.0 to 29.17, which contained the adenovirus homologs of IVa2 protein and the DNA replication proteins, precursor of terminal protein and DNA polymerase proteins. Analysis of the sequence for cis-acting elements suggests that transcripts of DNA polymerase and precursor of terminal protein are 3' co-terminal. In addition, this region also contains major late promoter sequence. The sequence to the left of IVa2 contains the ORF of pIX with a potential TATA box immediately upstream and two polyadenylation consensus signals immediately downstream of the ORF.

A retrospective analysis of early excision and skin grafting from 1993-1995.

Early excision and skin grafting (EEG) is an established and accepted procedure for deep burn management. This is a retrospective analysis of 100 patients treated with early excision and grafting in burns of all types and up to 65 per cent TBSA. Excisional surgery was performed in the period from day 2 to day 7 post-burn in patients who did not have significant infection (<10(5)) in one to two stages. The average mortality rate in all age groups and including smoke inhalation injury in our unit was 43.4 per cent. The mortality in the operated group was 10.2 per cent. The main causes of mortality were smoke inhalation injury, septicaemia (probably originating from the non-excised tissue) and extensive burn injury. The functional and aesthetic outcome in EEG patients was far superior in comparison with the conventional method of treatment.

Nucleotide sequence, genome organization, and transcription map of bovine adenovirus type 3.

The complete DNA sequence of bovine adenovirus type 3 is reported here. The size of the genome is 34,446 bp in length with a G+C content of 54%. All the genes of the early and late regions are present in the expected locations of the genome. However, the late-region genes are organized into seven families, instead of five as they are in human adenovirus type 2. The deduced amino acid sequences of open reading frames (ORFs) in the late regions and early region 2 (E2) and for IVa2 show higher degrees of homology, whereas the predicted amino acid sequences of ORFs in the E1, E3, and E4 regions and the pIX, fiber, and 33,000-molecular-weight nonstructural proteins show little or no homology with the corresponding proteins of other adenoviruses. In addition, the penton base protein lacks the integrin binding motif, RGD, but has an LDV motif instead of an MDV motif. Interestingly, as in other animal adenoviruses, the virus-associated RNA genes appear to be absent from their usual location. Sequence analysis of cDNA clones representing the early- and late-region genes identified splice acceptor and splice donor sites, polyadenylation signals and polyadenylation sites, and tripartite leader sequences.

Neurotrophins and their receptors in nerve injury and repair.

Cytokines are a heterogenous group of polypeptide mediators that have been associated with activation of numerous functions, including the immune system and inflammatory responses. The cytokine families include, but are not limited to, interleukins (IL-I alpha, IL-I beta, ILIra and IL-2-IL-15), chemokines (IL-8/ NAP-I, NAP-2, MIP-I alpha and beta, MCAF/MCP-1, MGSA and RANTES), tumor necrosis factors (TNF-alpha and TNF-beta), interferons (INF-alpha, beta and gamma), colony stimulating factors (G-CSF, M-CSF, GM-CSF, IL-3 and some of the other ILs), growth factors (EGF, FGF, PDGF, TGF alpha, TGF beta and ECGF), neuropoietins (LIF, CNTF, OM and IL-6), and neurotrophins (BDNF, NGF, NT-3-NT-6 and GDNF). The neurotrophins represent a family of survival and differentiation factors that exert profound effects in the central and peripheral nervous system (PNS). The neurotrophins are currently under investigation as therapeutic agents for the treatment of neurodegenerative disorders and nerve injury either individually or in combination with other trophic factors such as ciliary neurotrophic factor (CNTF) or fibroblast growth factor (FGF). Responsiveness of neurons to a given neurotrophin is governed by the expression of two classes of cell surface receptor. For nerve growth factor (NGF), these are p75NTR (p75) and p140trk (referred to as trk or trkA), which binds both BDNF and neurotrophin (NT)-4/5, and trkC receptor, which binds only NT-3. After binding ligand, the neurotrophin-receptor complex is internalized and retrogradely transported in the axon to the soma. Both receptors undergo ligand-induced dimerization, which activates multiple signal transduction pathways. These include the ras-dependent pathway utilized by trk to mediate neurotrophin effects such as survival and differentiation. Indeed, cellular diversity in the nervous system evolves from the concerted processes of cell proliferation, differentiation, migration, survival, and synapse formation. Neural adhesion and extracellular matrix molecules have been shown to play crucial roles in axonal migration, guidance, and growth cone targeting. Proinflammatory cytokines, released by activated macrophages and monocytes during infection, can act on neural targets that control thermogenesis, behavior, and mood. In addition to induction of fever, cytokines induce other biological functions associated with the acute phase response, including hypophagia and sleep. Cytokine production has been detected within the central nervous system as a result of brain injury, following stab wound to the brain, during viral and bacterial infections (AIDS and meningitis), and in neurodegenerative processes (multiple sclerosis and Alzheimer's disease). Novel cytokine therapies, such as anticytokine antibodies or specific receptor antagonists acting on the cytokine network may provide an optimistic feature for treatment of multiple sclerosis and other diseases in which cytokines have been implicated.

Oxidative stress and antioxidant therapy in Parkinson's disease.

Parkinson's disease, known also as striatal dopamine deficiency syndrome, is a degenerative disorder of the central nervous system characterized by akinesia, muscular rigidity, tremor at rest, and postural abnormalities. In early stages of parkinsonism, there appears to be a compensatory increase in the number of dopamine receptors to accommodate the initial loss of dopamine neurons. As the disease progresses, the number of dopamine receptors decreases, apparently due to the concomitant degeneration of dopamine target sites on striatal neurons. The loss of dopaminergic neurons in Parkinson's disease results in enhanced metabolism of dopamine, augmenting the formation of H2O2, thus leading to generation of highly neurotoxic hydroxyl radicals (OH.). The generation of free radicals can also be produced by 6-hydroxydopamine or MPTP which destroys striatal dopaminergic neurons causing parkinsonism in experimental animals as well as human beings. Studies of the substantia nigra after death in Parkinson's disease have suggested the presence of oxidative stress and depletion of reduced glutathione; a high level of total iron with reduced level of ferritin; and deficiency of mitochondrial complex I. New approaches designed to attenuate the effects of oxidative stress and to provide neuroprotection of striatal dopaminergic neurons in Parkinson's disease include blocking dopamine transporter by mazindol, blocking NMDA receptors by dizocilpine maleate, enhancing the survival of neurons by giving brain-derived neurotrophic factors, providing antioxidants such as vitamin E, or inhibiting monoamine oxidase B (MAO-B) by selegiline. Among all of these experimental therapeutic refinements, the use of selegiline has been most successful in that it has been shown that selegiline may have a neurotrophic factor-like action rescuing striatal neurons and prolonging the survival of patients with Parkinson's disease.

Molecular studies of the acute infection, latency and reactivation of equine herpesvirus-1 (EHV-1) in the mouse model.

The murine intranasal (i.n.) infection model was used to study the molecular distribution of equine herpesvirus-1 (EHV-1) during acute infection, latency and following a reactivation stimulus. After inoculation, infectious virus was detected in lungs, nasal turbinates, brains and olfactory bulbs during the acute phase. A nested PCR (nPCR) readily detected virus in these tissues and, in addition, virus was detected in spleens and (in the second round of nPCR) in peripheral blood mononuclear cells (PBMC). A digoxigenin-labelled in situ hybridization probe detected EHV-1 DNA in bronchiolar and vascular endothelium in the lungs and in and around germinal centres in the spleens. One month later, although infectious virus was absent from all tissues, the trigeminal ganglia, olfactory bulb and PBMC remained positive for virus DNA although this was detected only on the second round of nPCR. Furthermore, in situ hybridization, using either DNA or RNA probes, suggested that little or no transcription of virus occurred in neural tissues during the 'latent phase'. Following a reactivation stimulus, infectious virus was not isolated from any tissues, however, EHV-1 DNA was detected on the first round of nPCR in olfactory bulb, trigeminal ganglia and PBMC. This suggested a quantitative increase in EHV-1 DNA occurred following reactivation stimulus. The significance of these results is discussed in relation to the molecular state of EHV-1 in different tissues at various stages of infection and the validity of the murine model for studying latency and reactivation of EHV-1 in the horses.

The detection of latency-associated transcripts of equine herpesvirus 1 in ganglionic neurons.

Neural tissues from specific pathogen-free ponies that had been experimentally infected with equine herpesvirus 1 (EHV-1) were analysed by in situ hybridization. Digoxigenin-labelled EHV-1 BamHI fragments spanning almost the entire EHV-1 genome were hybridized to RNA in tissue sections from latently infected trigeminal ganglia. The BamHI E fragment detected EHV-1 RNA antisense to gene 63 (HSV-1 homologue ICP0) in a small number of neurons. Sixteen other BamHI fragments gave negative results in 20 sections tested with each fragment. Latency associated transcripts (LATs) were localized to the neuronal nuclei. EHV-1 nucleotide sequence data in the region reveals the presence of a putative EHV-1 LAT promoter that shares a similar motifs with the HSV-1 LAT promoter, including the LAT promoter-binding factor, and may have a role in EHV-1 LAT expression.