Multiple steps of prion strain adaptation to a new host.

The transmission of prions across species is a critical aspect of their dissemination among mammalian hosts, including humans. This process often necessitates strain adaptation. In this study, we sought to investigate the mechanisms underlying prion adaptation while mitigating biases associated with the history of cross-species transmission of natural prion strains. To achieve this, we utilized the synthetic hamster prion strain S05. Propagation of S05 using mouse PrPC in Protein Misfolding Cyclic Amplification did not immediately overcome the species barrier. This finding underscores the involvement of factors beyond disparities in primary protein structures. Subsequently, we performed five serial passages to stabilize the incubation time to disease in mice. The levels of PrPSc increased with each passage, reaching a maximum at the third passage, and declining thereafter. This suggests that only the initial stage of adaptation is primarily driven by an acceleration in PrPSc replication. During the protracted adaptation to a new host, we observed significant alterations in the glycoform ratio and sialylation status of PrPSc N-glycans. These changes support the notion that qualitative modifications in PrPSc contribute to a more rapid disease progression. Furthermore, consistent with the decline in sialylation, a cue for "eat me" signaling, the newly adapted strain exhibited preferential colocalization with microglia. In contrast to PrPSc dynamics, the intensity of microglia activation continued to increase after the third passage in the new host. In summary, our study elucidates that the adaptation of a prion strain to a new host is a multi-step process driven by several factors.

Multiple steps of prion strain adaptation to a new host.

The transmission of prions across species is a critical aspect of their dissemination among mammalian hosts, including humans. This process often necessitates strain adaptation. In this study, we sought to investigate the mechanisms underlying prion adaptation while mitigating biases associated with the history of cross-species transmission of natural prion strains. To achieve this, we utilized the synthetic hamster prion strain S05. Propagation of S05 using mouse PrPC in Protein Misfolding Cyclic Amplification did not immediately overcome the species barrier. This finding underscores the involvement of factors beyond disparities in primary protein structures. Subsequently, we performed five serial passages to stabilize the incubation time to disease in mice. The levels of PrPSc increased with each passage, reaching a maximum at the third passage, and declining thereafter. This suggests that only the initial stage of adaptation is primarily driven by an acceleration in PrPSc replication. During the protracted adaptation to a new host, we observed significant alterations in the glycoform ratio and sialylation status of PrPSc N-glycans. These changes support the notion that qualitative modifications in PrPSc contribute to a more rapid disease progression. Furthermore, consistent with the decline in sialylation, a cue for "eat me" signaling, the newly adapted strain exhibited preferential colocalization with microglia. In contrast to PrPSc dynamics, the intensity of microglia activation continued to increase after the third passage in the new host. In summary, our study elucidates that the adaptation of a prion strain to a new host is a multi-step process driven by several factors.

Reactive astrocytes associated with prion disease impair the blood brain barrier.

BACKGROUND: Impairment of the blood-brain barrier (BBB) is considered to be a common feature among neurodegenerative diseases, including Alzheimer's, Parkinson's and prion diseases. In prion disease, increased BBB permeability was reported 40 years ago, yet the mechanisms behind the loss of BBB integrity have never been explored. Recently, we showed that reactive astrocytes associated with prion diseases are neurotoxic. The current work examines the potential link between astrocyte reactivity and BBB breakdown. RESULTS: In prion-infected mice, the loss of BBB integrity and aberrant localization of aquaporin 4 (AQP4), a sign of retraction of astrocytic endfeet from blood vessels, were noticeable prior to disease onset. Gaps in cell-to-cell junctions along blood vessels, together with downregulation of Occludin, Claudin-5 and VE-cadherin, which constitute tight and adherens junctions, suggested that loss of BBB integrity is linked with degeneration of vascular endothelial cells. In contrast to cells isolated from non-infected adult mice, endothelial cells originating from prion-infected mice displayed disease-associated changes, including lower levels of Occludin, Claudin-5 and VE-cadherin expression, impaired tight and adherens junctions, and reduced trans-endothelial electrical resistance (TEER). Endothelial cells isolated from non-infected mice, when co-cultured with reactive astrocytes isolated from prion-infected animals or treated with media conditioned by the reactive astrocytes, developed the disease-associated phenotype observed in the endothelial cells from prion-infected mice. Reactive astrocytes were found to produce high levels of secreted IL-6, and treatment of endothelial monolayers originating from non-infected animals with recombinant IL-6 alone reduced their TEER. Remarkably, treatment with extracellular vesicles produced by normal astrocytes partially reversed the disease phenotype of endothelial cells isolated from prion-infected animals. CONCLUSIONS: To our knowledge, the current work is the first to illustrate early BBB breakdown in prion disease and to document that reactive astrocytes associated with prion disease are detrimental to BBB integrity. Moreover, our findings suggest that the harmful effects are linked to proinflammatory factors secreted by reactive astrocytes.

Reactive astrocytes associated with prion disease impair the blood brain barrier.

Background: Impairment of the blood-brain barrier (BBB) is considered to be a common feature among neurodegenerative diseases, including Alzheimer's, Parkinson's and prion diseases. In prion disease, increased BBB permeability was reported 40 years ago, yet the mechanisms behind the loss of BBB integrity have never been explored. Recently, we showed that reactive astrocytes associated with prion diseases are neurotoxic. The current work examines the potential link between astrocyte reactivity and BBB breakdown. Results: In prion-infected mice, the loss of BBB integrity and aberrant localization of aquaporin 4 (AQP4), a sign of retraction of astrocytic endfeet from blood vessels, were noticeable prior to disease onset. Gaps in cell-to-cell junctions along blood vessels, together with downregulation of Occludin, Claudin-5 and VE-cadherin, which constitute tight and adherens junctions, suggested that loss of BBB integrity is linked with degeneration of vascular endothelial cells. In contrast to cells isolated from non-infected adult mice, endothelial cells originating from prion-infected mice displayed disease-associated changes, including lower levels of Occludin, Claudin-5 and VE-cadherin expression, impaired tight and adherens junctions, and reduced trans-endothelial electrical resistance (TEER). Endothelial cells isolated from non-infected mice, when co-cultured with reactive astrocytes isolated from prion-infected animals or treated with media conditioned by the reactive astrocytes, developed the disease-associated phenotype observed in the endothelial cells from prion-infected mice. Reactive astrocytes were found to produce high levels of secreted IL-6, and treatment of endothelial monolayers originating from non-infected animals with recombinant IL-6 alone reduced their TEER. Remarkably, treatment with extracellular vesicles produced by normal astrocytes partially reversed the disease phenotype of endothelial cells isolated from prion-infected animals. Conclusions: To our knowledge, the current work is the first to illustrate early BBB breakdown in prion disease and to document that reactive astrocytes associated with prion disease are detrimental to BBB integrity. Moreover, our findings suggest that the harmful effects are linked to proinflammatory factors secreted by reactive astrocytes.

Aß plaques do not protect against HSV-1 infection in a mouse model of familial Alzheimer's disease, and HSV-1 does not induce Aß pathology in a model of late onset Alzheimer's disease.

The possibility that the etiology of late onset Alzheimer's disease is linked to viral infections of the CNS has been actively debated in recent years. According to the antiviral protection hypothesis, viral pathogens trigger aggregation of Aß peptides that are produced as a defense mechanism in response to infection to entrap and neutralize pathogens. To test the causative relationship between viral infection and Aß aggregation, the current study examined whether Aß plaques protect the mouse brain against Herpes Simplex Virus 1 (HSV-1) infection introduced via a physiological route and whether HSV-1 infection triggers formation of Aß plaques in a mouse model of late-onset AD that does not develop Aß pathology spontaneously. In aged 5XFAD mice infected via eye scarification, high density of Aß aggregates did not improve survival time or rate when compared with wild type controls. In 5XFADs, viral replication sites were found in brain areas with a high density of extracellular Aß deposits, however, no association between HSV-1 and Aß aggregates could be found. To test whether HSV-1 triggers Aß aggregation in a mouse model that lacks spontaneous Aß pathology, 13-month-old hAß/APOE4/Trem2*R47H mice were infected with HSV-1 via eye scarification with the McKrae HSV-1 strain, intracranial inoculation with McKrae, intracranial inoculation after priming with LPS for 6 weeks, or intracranial inoculation with high doses of McKrae or 17syn + strains that represent different degrees of neurovirulence. No signs of Aß aggregation were found in any of the experimental groups. Instead, extensive infiltration of peripheral leukocytes was observed during the acute stage of HSV-1 infection, and phagocytic activity of myeloid cells was identified as the primary defense mechanism against HSV-1. The current results argue against a direct causative relationship between HSV-1 infection and Aß pathology.

The Roles of USG and NCCT in the Diagnosis of Acute Appendicitis: A Study in a Tertiary Care Center in North Eastern India.

Background: Acute appendicitis is a common cause of hospital admission and emergency laparotomy among children and young adults. Although the diagnosis is clinical, the use of radiological imaging has emerged over the past decades. Its principal use is as a problem-solving tool in equivocal cases. Owing to the increased use of imaging in the last few years, the negative appendicectomy rate has dropped significantly. In this prospective observational study, we compared the diagnostic accuracy of Ultrasonography and Non-Contrast Computed Tomography. Method: One hundred and eighteen patients with clinically suspected appendicitis followed a designed protocol. Patients underwent appendicectomy after a first performed positive ultrasonography or after a positive Non-Contrast Computed Tomography when Ultrasonography was equivocal or nonspecific. When any other diagnosis was apparent in either imaging modality which could explain the symptomatology in the patient, they were considered negative for acute appendicitis and treated accordingly. Results: The respective sensitivity, specificity, and accuracy for Ultrasonography, Non-Contrast Computed Tomography, and the whole diagnostic pathway for the diagnosis of acute appendicitis were 70.73%,80.83%, and 78.54; 100%,100%,100%, and 83.6%; and 100%,83.33% and 94.92%. Conclusion: Using Ultrasonography as the first-line diagnostic tool and Non-Contrast Computed Tomography as a complementary second-line diagnostic tool, appendicitis can be diagnosed with high accuracy and the negative laparotomy rate can be brought down significantly without any increase in the risk of complications. Computed Tomography is superior to Ultrasonography for the diagnosis of acute appendicitis.

Treatment of long bone infection by a biodegradable bone cement releasing antibiotics in human.

Repair of methicillin-resistant Staphylococcal (MRSA) chronic osteomyelitis and resulting bone defect is one of the major challenges in orthopaedics. Previous study has shown the effectiveness of antibiotic loaded biodegradable composite bone cement with in vitro tests and in the treatment of experimental osteomyelitis. The cement is composed of poly(lactide-co-glycolide) encapsulated antibiotic-biphasic calcium phosphate granule complex and additive antibiotic powder in gypsum binder. In this study, the cement was studied further to evaluate its in vitro biological properties (cytocompatibility, platelet activation), anti-infective, and bone regenerative potential in comparison to poly(methyl methacrylate) (PMMA) cement and parenteral therapy in 43 patients (age 5-57 years) with chronic MRSA osteomyelitis by analyzing the results of histopathology, radiographs, magnetic resonance imaging, scanning electron microscopy, and serum drug concentrations for 1 year. The composite cement showed superior cytocompatibility and coagulant activity compared to PMMA cement. Moreover, the results of different postoperative clinical and radiological examinations also proved the supremacy of composite cement over the other treatment modalities in terms of success rate, faster sepsis control and bone regeneration. Low serum antibiotic concentrations and normal serum calcium levels indicate that the calcium-rich composite cement is safe for application in human. Therefore, we conclude that the composite bone cement is a promising candidate for the treatment of chronic osteomyelitis.

Alzheimer's disease-associated ß-amyloid does not protect against herpes simplex virus 1 infection in the mouse brain.

Alzheimer's disease (AD) is a devastating fatal neurodegenerative disease. An alternative to the amyloid cascade hypothesis is that a viral infection is key to the etiology of late-onset AD, with ß-amyloid (Aß) peptides playing a protective role. In the current study, young 5XFAD mice that overexpress mutant human amyloid precursor protein with the Swedish, Florida, and London familial AD mutations were infected with one of two strains of herpes simplex virus 1 (HSV-1), 17syn+ and McKrae, at three different doses. Contrary to previous work, 5XFAD genotype failed to protect mice against HSV-1 infection. The region- and cell-specific tropisms of HSV-1 were not affected by the 5XFAD genotype, indicating that host-pathogen interactions were not altered. Seven- to ten-month-old 5XFAD animals in which extracellular Aß aggregates were abundant showed slightly better survival rate relative to their wild-type (WT) littermates, although the difference was not statistically significant. In these 5XFAD mice, HSV-1 replication centers were partially excluded from the brain areas with high densities of Aß aggregates. Aß aggregates were free of HSV-1 viral particles, and the limited viral invasion to areas with a high density of Aß aggregates was attributed to phagocytic activity of reactive microglia. In the oldest mice (12-15 months old), the survival rate did not differ between 5XFAD and WT littermates. While the current study questions the antiviral role of Aß, it neither supports nor refutes the viral etiology hypothesis of late-onset AD.

Joint scores in hemophilic arthropathy in children: Developing country perspectives.

OBJECTIVE: Hemophilia is a common X-linked recessive coagulopathy causing recurrent bleeding into the synovial joints and results in articular and periarticular abnormalities. To our knowledge, this is the first comprehensive study aimed at studying the clinico-radiological joint score evaluation in hemophilic arthropathy in children from a developing country and its possible impact on the quality of life. METHODS: In this hospital-based, prospective, descriptive study, all children presenting to the pediatric rheumatology clinic were studied. The joint physical examination was scored using the Hemophilia Joint Health Score 2.1 (HJHS 2.1). The patients were then subjected to imaging of the most affected joint using ultrasonography (USG) and magnetic resonance imaging (MRI). Detailed USG and MRI radiological evaluation was recorded in the predesigned proforma using the Hemophilia Early Arthropathy Detection with Ultra Sound (HEAD-US) score and MRI DENVER score. The physical quality of life as per Functional Independence Score in Hemophilia (FISH) was noted. The clinical, radiological, and functional scores were analyzed with an appropriate statistical measure. RESULTS: The mean age at presentation was 7.4 years (interquartile range 4.9-10), with the knee being the most common joint involved. All of the USG score, MRI score, and FISH score have a significant correlation (p<0.05), with the HJHS 2.1 score with correlation coefficients of 0.7086, -0.8916, and 0.8607, respectively. USG and MRI had a correlation coefficient of -0.7145 and -0.8326 with FISH, respectively. CONCLUSION: The degree of association between HJHS 2.1 score was found to be maximum with HEAD-US score, whereas a negative correlation was seen evaluating FISH score with both HEAD-US and MRI DENVER scores. Use of these scores, specifically HEAD-US score, will result in consistent assessment of hemophilic joints, optimizing the management of the destructive changes.

Refractory Seizure in Childhood: Dyke-Davidoff-Masson Syndrome Revisited.

Dyke-Davidoff-Masson syndrome (DDMS) is a rare disorder characterized by recurrent seizures, facial asymmetry, contralateral hemiplegia, radiologic features of cerebral hemiatrophy, and ipsilateral compensatory hypertrophy of the skull bone and sinuses. We describe three cases of children with DDMS, who initially presented with refractory seizure to the pediatric department of North Bengal Medical College and Hospital, India. In each case, the clinical features noted along with computed tomography or magnetic resonance imaging helped confirm the diagnosis of DDMS. DDMS should be considered as a differential diagnosis of refractory seizures in children. We seek to emphasize the importance of thorough clinical and neuroimaging workup of seizure disorder in children for the proper management of the condition.

Osteomeatal Complex: A Study of Its Anatomical Variation Among Patients Attending North Bengal Medical College and Hospital.

Lateral nasal wall of each nasal cavity provides the final common pathway of drainage of the mucociliary clearance of frontal, maxillary and anterior ethmoidal air cells. Anatomical variants like concha bullosa, Haller cells, agger nasi cells, enlarged bulla ethmoidalis may obstruct the mucociliary clearance through osteomeatal complex and cause rhino sinusitis. The objectives were to find out the anatomical variation of osteomeatal complex and its dimensions when present. The present study was a descriptive, hospital based cross sectional study carried out in the outpatient departments of North Bengal Medical College and Hospital, West Bengal, India, among patients aged 15 years and above. Coronal CT scan of paranasal sinus and orbit region was done. Data was collected with the help of semi structured predesigned and pretested questionnaire. Of the 44 study patients, 15.9 % had concha bullosa, 11.36 % had paradoxical middle concha, 27.3 % had Haller cell, 18.2 % had agger nasi cell. Lateral attachment and medial free margin of uncinate process were also measured in respect to medial body line. 77.3, 59.1 and 47.7 % had sneezing, rhinorrhoea and headache respectively. The harmony of mucociliary clearance and obstruction free osteomeatal complex is the key factor for ventilation and drainage of maxillary, frontal and anterior ethmoidal air cells.

High temperature-induced sterility in the female Nile tilapia, Oreochromis niloticus.

High temperature treatments induce germ cell loss in gonads of vertebrate animals, including fish. It could be a reliable source for induction of sterility if the treatments led to a permanent loss of germ cells. Here we report that heat treatment at 37 °C for 45-60 days caused a complete loss of germ cells in female Nile tilapia, Oreochromis niloticus, and that sterility was achieved in fish at all stages of their life cycle. Unlike previous observations, germ cells did not repopulate even after returning them to the water at control conditions suggesting permanent depletion of germ cells. Gonadal somatic cells immunopositive for 3ß-hydroxysteroid dehydrogenase (3ß-HSD) were clustered at one end of the germ cell depleted gonads close to the blood vessel. Serum level of testosterone, 11-ketotestosterone, and 17ß-estradiol was significantly decreased in sterile fish compared to control. Body weight of sterile fish was higher than control fish at the end of experiment. Our observations of increased growth and permanent sterilization in the high temperature-treated fish suggest that this method could be an appropriate and eco-friendly tool for inducing sterility in fish with a higher thermal tolerance.

Identification, characterization and feeding response of peptide YYb (PYYb) gene in grass carp (Ctenopharyngodon idellus).

The peptide YYb (PYYb) is a fish-specific peptide belonging to the neuropeptide Y (NPY) family. In the present study, the full-length cDNA sequence and genomic structure of PYYb (gcPYYb) from Ctenopharyngodon idellus have been isolated and characterized. The gcPYYb gene consists of three exons interspaced by two introns, opposing to the typical architecture of most NPY-family genes as well as its paralogs. Alignment of deduced amino acid sequence indicates that the fish PYYb is more variable compared to NPY and PYY as shown by more residue changes in teleosts lineage, suggesting mild selective pressure imposed on the peptide. Real-time quantitative PCR analysis shows the gcPYYb mRNA in developing larvae is increased during the mixed endo- and exogenous-feeding period and is widely distributed throughout the intestine of fully grown individuals. Following a single meal, the gcPYYb mRNA in foregut is increased at 3 h post-feeding and subsequently decreased before the foregut contents are cleared. These results suggest that the gcPYYb has an important role in the early life stages of grass carp and is involved in food intake by transmitting feeding-related signals.

A case of bilaterally elongated styloid process differing in osseo-morphology.

In a cranium of a male cadaver styloid processes have been found to be unusually long with different lengths. Elongation of styloid process involvethe entire 'Stylohyoid complex/chain', though not rare as reported in earlier literatures, but the osseomorphological and radiological analysis of present case brings out a unique variety as its described in the article.

Molecular cloning, expression analysis, and potential food intake attenuation effect of peptide YY in grass carp (Ctenopharyngodon idellus).

The peptide YY (PYY) is a 36 amino acid peptide involved in the food intake control in vertebrates. We have cloned and characterized a PYY gene from grass carp Ctenopharyngodon idellus. The full-length cDNA encodes a precursor protein of grass carp PYY (gcPYY) that consists of a putative 28-amino acid signal peptide, a 36-amino acid mature peptide, an amidation-proteolytic site, and a 30-amino acid carboxy-terminal extension. The gcPYY gene is comprised of 4 exons interspaced by 3 introns as seen in PYYs from other species. Amino acid alignment and gene structure comparison indicate that the structure of PYY is well preserved throughout vertebrate phylogeny. The tissue distribution and postprandial changes in gcPYY mRNA expression were evaluated by real-time PCR, which showed that the gcPYY is expressed abundantly in the central nervous system, with significantly increased expression following a single meal. During embryogenesis, the presence of gcPYY mRNA was detected in early developing embryos, and high expression levels were observed when most larvae completed their switch from endogenous nourishment to exogenous feeding. Reduced food intake by juveniles during a single meal after giving perpheral injection of gcPYY1-36 suggests a potentially important role of PYY in the food intake attenuation in grass carp.

Molecular cloning, characterization and immunological response analysis of Toll-like receptor 21 (TLR21) gene in grass carp, Ctenopharyngodon idella.

TLR21, a non-mammalian Toll-like receptor, has been recently identified in fishes, frogs and birds. In the present study, the full-length cDNA sequence of TLR21 (CiTLR21) from Ctenopharyngodon idella has been isolated and characterized. The CiTLR21 full-length cDNA sequence consists of 3578bp, with an open reading frame (ORF) of 2958bp encoding 985 amino acid residues. The putative CiTLR21 protein contains a signal peptide sequence, 17 leucine-rich (LRR) motifs, a transmembrane region and a Toll/interleukin-1 receptor (TIR) domain. The CiTLR21 gene is expressed in a wide range of tissues with the highest expression in skin. Upon induction by Aquareovirus, CiTLR21 expression is significantly down-regulated in liver and spleen, whereas is significantly up-regulated in liver and spleen after Aeromonas hydrophila infection. These results suggest that CiTLR21 plays an important role in Aquareovirus and A. hydrophila-related diseases. This work may provide the basis for further investigations into the immune system of grass carp and other teleost fishes.

Identification of TNF13b (BAFF) gene from grass carp (Ctenopharyngodon idella) and its immune response to bacteria and virus.

The TNF superfamily B cell activating factor (BAFF) is a central cytokine in several diseases. A BAFF gene has been cloned from grass carp (Ctenopharyngodon idella), analyzed its structure, and investigated its expression pattern in various tissues after Aeromonas hydrophila and Aquareovirus infection. The open reading frame of grass carp BAFF (gcBAFF) consists of 804 bases encoding 267 amino acids. Phylogenetic analysis shows the gcBAFF is most closely related to other teleost BAFFs with the highest similarity to zebrafish. RT-PCR analysis shows the gcBAFF transcript is expressed in a wide range of tissues with the highest expression in skin and spleen. Upon induction by A. hydrophila and Aquareovirus, its expression is significantly up-regulated in gill, liver, kidney, spleen and skin as compared to PBS injected fish. The association of increased BAFF expression after bacterial and viral infections suggests that it plays a potentially important role in immune system of fish.

Growth hormone deficiency in fanconis anemia.

Two cases of Fanconis anemia (FA) are reported here. Case 1 presented with hypoplastic anemia and skeletal abnormality. Case 2, his older brother had stunted growth, investigations detected growth hormone deficiency and mild hematological derangement without other endocrine abnormality. FA was confirmed by positive chromosomal breakages studies in both cases.

Childhood steroid-responsive ophthalmoplegic migraine.

Ophthalmoplegic migraine (OM) is characterized by recurrent attacks of headache with paresis of ocular cranial nerves. Previously, it was classified as a variant of migraine, but recently, International Headache Classification (IHCD-II) has reclassified OM to the category of neuralgia. Presently, OM is considered a type of recurrent demyelinating cranial neuropathy. We report an adolescent girl with OM, who had been treated with steroid and showed dramatic improvement.

Sirenomelia.

Sirenomelia is a rare malformation of caudal part of embryo. It is characterised by complete or partial fusion of the legs into a single lower limb. Abnormalities of the kidneys, large intestines and genitalia are common. Sirenomelia cases have only one umbilical artery and one vein. Upper body birth defects are rare and include abnormalities of heart, lungs, arms, spine and brain. Here we report a case of sirenomelia with uncommon upper body birth defects involving right forearm and hand, and the rib cage. Vascular steal phenomenon cannot explain the upper body birth defects.