Mechanotransduction, PROX1, and FOXC2 cooperate to control connexin37 and calcineurin during lymphatic-valve formation.

Lymphatic valves are essential for efficient lymphatic transport, but the mechanisms of early lymphatic-valve morphogenesis and the role of biomechanical forces are not well understood. We found that the transcription factors PROX1 and FOXC2, highly expressed from the onset of valve formation, mediate segregation of lymphatic-valve-forming cells and cell mechanosensory responses to shear stress in vitro. Mechanistically, PROX1, FOXC2, and flow coordinately control expression of the gap junction protein connexin37 and activation of calcineurin/NFAT signaling. Connexin37 and calcineurin are required for the assembly and delimitation of lymphatic valve territory during development and for its postnatal maintenance. We propose a model in which regionally increased levels/activation states of transcription factors cooperate with mechanotransduction to induce a discrete cell-signaling pattern and morphogenetic event, such as formation of lymphatic valves. Our results also provide molecular insights into the role of endothelial cell identity in the regulation of vascular mechanotransduction.

Lack of association between connexin40 polymorphisms and coronary artery disease.

OBJECTIVE: Cx40 is a gap junction protein important for cell-cell communication in the endothelium. Polymorphisms in the promoter region of the human Cx40 gene, -44G>A and +71A>G, were shown to reduce Cx40 transcription by half. As mice with an endothelial-specific deletion of Cx40 are more susceptible to atherosclerosis, this study was designed to discover a correlation between these polymorphisms and atherosclerosis in European populations.¦METHODS AND RESULTS: 803 patients referred to the Geneva University Hospitals for elective coronary angiography were divided according to the number of significantly stenosed vessels (from 0 to 3) and were genotyped for the Cx40 polymorphisms. Genotype distribution in the control group was -44GG/+71AA=59.8%, -44AG/+71AG=35.1% and -44AA/+71GG=5.2%. Surprisingly, this distribution was similar in the CAD group, with -44GG/+71AA=58.5%, -44AG/+71AG=37.6% and -44AA/+71GG=3.8% (p=0.67). Moreover, no significant association between histological carotid plaque composition of culprit lesions and Cx40 polymorphisms could be detected in 583 Dutch patients of the Athero-Express study.¦CONCLUSIONS: Despite a clear antiatherogenic role of Cx40 in mice, our study could not detect an association of Cx40 promoter polymorphisms and CAD in human. Moreover, a correlation with atherosclerotic plaque stability or hypertension could not be demonstrated either. Connexin polymorphisms affecting channel function may be of greater importance for cardiovascular disease than polymorphisms affecting the expression level of the protein.

Connexin 37 limits thrombus propensity by downregulating platelet reactivity.

Background- Formation of platelet plug initiates hemostasis after vascular injury and triggers thrombosis in ischemic disease. However, the mechanisms leading to the formation of a stable thrombus are poorly understood. Connexins comprise a family of proteins that form gap junctions enabling intercellular coordination of tissue activity, a process termed gap junctional intercellular communication. Methods and Results- In the present study, we show that megakaryocytes and platelets express connexin 37 (Cx37). Deletion of the Cx37 gene in mice shortens bleeding time and increases thrombus propensity. Aggregation is increased in murine Cx37(-/-) platelets or in murine Cx37(+/+) and human platelets treated with gap junction blockers. Intracellular microinjection of neurobiotin, a Cx37-permeant tracer, revealed gap junctional intercellular communication in platelet aggregates, which was impaired in Cx37(-/-) platelets and in human platelets exposed to gap junction blockers. Finally, healthy subjects homozygous for Cx37-1019C, a prognostic marker for atherosclerosis, display increased platelet responses compared with subjects carrying the Cx37-1019T allele. Expression of these polymorphic channels in communication-deficient cells revealed a decreased permeability of Cx37-1019C channels for neurobiotin. Conclusions- We propose that the establishment of gap junctional communication between Cx37-expressing platelets provides a mechanism to limit thrombus propensity. To our knowledge, these data provide the first evidence incriminating gap junctions in the pathogenesis of thrombosis.

Trichurid nematodes in ring-necked pheasants from backyard flocks of the State of Rio de Janeiro, Brazil: frequency and pathology

The present investigation is related to the frequency of infection and to the gross and microscopic lesions associated to the presence of trichurid worms in 50 ring-necked pheasants (Phasianus colchicus) from backyard flocks in the state of Rio de Janeiro, Brazil. In the investigated birds, the overall infection rate was of 74%, with the presence of Eucoleus perforans with 72% of prevalence and 21.2 of mean intensity, in the esophageal and crop mucosa and rarely in the junction of the proventriculus and esophagus, E. annulatus with 2% and 3 in the crop mucosa, Capillaria phasianina, with 12% and 4.3 in the cecum and small intestine and Baruscapillaria obsignata, for the first time referred in this host, with 2% and 1 in the small intestine. Clinical signs were absent. The gross lesions observed in the crop and esophagus of 14 (38.9%) pheasants parasitized with E. perforans were thickening, small nodules, congestion, and petechial haemorrhages in the mucosa. These birds presented a mean infection of 37.5 and a range of infection of 10-82. The microscopic lesions revealed chronic esophagitis with diffuse inflammatory process in the lamina propria characterized mostly by a mononuclear cell infiltrate and also with the presence of granulocytes. In the case of the parasitism of pheasants with C. phasianina, the gross lesions were absent; microscopic lesions were characterized by chronic typhlitis with mononuclear infiltrate. Gross and microscopic lesions were absent in the pheasants parasitized with E. annulatus and B. obsignata.

A New Large Deletion in the DFNB1 Locus Causes Nonsyndromic Hearing Loss.

Mutations in the GJB2 gene encoding the gap junction protein connexin 26 are responsible for up to 30% of all cases of autosomal recessive nonsyndromic hearing impairment (HI) with prelingual onset in most populations. The corresponding locus DFNB1, located on chromosome 13q11-q12, is also affected by three distinct deletions. These deletions extended distally to GJB2, which remains intact. We report a novel large deletion in DFNB1 observed in a patient presenting profound prelingual HI. This deletion was observed in trans to a GJB2 mutated allele carrying the p.Val84Met (V84M) mutation and was shown to be associated with hearing loss. The deletion caused a false homozygosity of V84M in the proband. Quantification of alleles by quantitative fluorescent multiplex PCR (QFM-PCR) enabled us to study the breakpoints of the deletion. The deleted segment extended through at least 920kb and removed the three connexin genes GJA3, GJB2 and GJB6. The distal breakpoint inside intron 2 of CRYL1 gene differed from the breakpoints of the known DFNB1 deletions. This case highlights the importance of screening for large deletions in molecular studies of GJB2.

Ascending aorta measurements as assessed by ECG-gated multi-detector computed tomography : a pilot study to etablish normative values for transcatheters therapies

Rapport de synthèse : Mesures de l'aorte ascendante par scanner synchronisé au rythme cardiaque: une étude pilote pour établir des valeurs normatives dans le cadre des futures thérapies par transcathéter. Objectif : L'objectif de cette étude est d'établir les valeurs morphométriques normatives de l'aorte ascendante à l'aide de l'angiographie par scanner synchronisé au rythme cardiaque, afin d'aider au développement des futurs traitements par transcathéter. Matériels et méthodes : Chez soixante-dix-sept patients (âgé de 22 à 83 ans, âge moyen: 54,7 ans), une angiographie par scanner synchronisé au rythme cardiaque a été réalisée pour évaluation des vaisseaux coronaires. Les examens ont été revus afin d'étudier l'anatomie de la chambre de chasse du ventricule gauche jusqu'au tronc brachio-céphalique droit. A l'aide de programmes de reconstructions multiplanaires et de segmentation automatique, différents diamètres et longueurs considérés comme importants pour les futurs traitements par transcathéter ont été mesurés. Les valeurs sont exprimées en moyennes, médianes, maximums, minimums, écart-types et en coefficients de variation. Les variations de diamètre de l'aorte ascendante durant le cycle cardiaque ont été aussi considérées. Résultats : Le diamètre moyen de la chambre de chasse du ventricule gauche était de 20.3+/-3.4 mm. Au niveau du sinus coronaire de l'aorte, il était de 34.2+/-4.1 mm et au niveau de la jonction sinotubulaire il était de 29.7+/-3.4 mm. Le diamètre moyen de l'aorte ascendante était de 32.7+/-3.8 mm. Le coefficient de variation de ces mesures variait de 12 à 17%. La distance moyenne entre l'insertion proximale des valvules aortiques et le départ du tronc brachio-céphalique droit était de 92.6+/-11.8 mm. La distance moyenne entre l'insertion proximale des valvules aortiques et l'origine de l'artère coronaire proximale était de 12.1+/-3.7 mm avec un coefficient de variation de 31%. La distance moyenne entre les deux ostia coronaires était de 7.2+/-3.1 mm avec un coefficient de variation de 43%. La longueur moyenne du petit arc de l'aorte ascendante entre l'artère coronaire gauche et le tronc brachio-céphalique droit était de 52.9+/-9.5 mm. La longueur moyenne de la continuité fibreuse entre la valve aortique et la valvule mitrale antérieure était de 14.6+/-3.3 mm avec un coefficient de variation de 23%. L'aire moyenne de la valve aortique était de 582.0+/-131.9 mm2. La variation du diamètre antéro-postérieur et transverse de l'aorte ascendante était respectivement de 8.4% et de 7.3%. Conclusion Il existe d'importantes variations inter-individuelles dans les mesures de l'aorte ascendante avec cependant des variations intra-individuelles faibles durant le cycle cardiaque. De ce fait, une approche personnalisée pour chaque patient est recommandée dans la confection des futures endoprothèses de l'aorte ascendante. Le scanner synchronisé au rythme cardiaque jouera un rôle prépondérant dans le bilan préthérapeutique. Abstract : The aim of this study was to provide an insight into normative values of the ascending aorta in regards to novel endovascular procedures using ECG-gated multi-detector CT angiography. Seventy-seven adult patients without ascending aortic abnormalities were evaluated. Measurements at relevant levels of the aortic root and ascending aorta were obtained. Diameter variations of the ascending aorta during cardiac cycle were also considered. Mean diameters (mm) were as follows: LV outflow tract 20.3+/-3.4, coronary sinus 34.2+/-4.1, sinotubular junction 29.7+-3.4 and mid ascending aorta 32.7+/-3.8 with coefficients of variation (CV) ranging from 12 to 17%. Mean distances (mm) were: from the plane passing through the proximal insertions of the aortic valve cusps to the right brachio-cephalic artery (BCA) 92.6111.8, from the plane passing through the proximal insertions of the aortic valve cusps to the proximal coronary ostium 12.1+/-3.7, and between both coronary ostia 7.2+/-3.1, minimal arc of the ascending aorta from left coronary ostium to right BCA 52.9 X9.5, and the fibrous continuity between the aortic valve and the anterior leaflet of the mitral valve 14.óf3.3, CV 13-43%. Mean aortic valve area was 582+-131.9 mm2. The variations of the antero-posterior and transverse diameters of the ascending aorta during the cardiac cycle were 8.4% and 7.3%, respectively. Results showed large inter-individual variations in diameters and distances but with limited intra-individual variations during the cardiac cycle. A personalized approach for planning endovascular devices must be considered.

Thyroid hormone enhances transected axonal regeneration and muscle reinnervation following rat sciatic nerve injury.

Improvement of nerve regeneration and functional recovery following nerve injury is a challenging problem in clinical research. We have already shown that following rat sciatic nerve transection, the local administration of triiodothyronine (T3) significantly increased the number and the myelination of regenerated axons. Functional recovery is a sum of the number of regenerated axons and reinnervation of denervated peripheral targets. In the present study, we investigated whether the increased number of regenerated axons by T3-treatment is linked to improved reinnervation of hind limb muscles. After transection of rat sciatic nerves, silicone or biodegradable nerve guides were implanted and filled with either T3 or phosphate buffer solution (PBS). Neuromuscular junctions (NMJs) were analyzed on gastrocnemius and plantar muscle sections stained with rhodamine alpha-bungarotoxin and neurofilament antibody. Four weeks after surgery, most end-plates (EPs) of operated limbs were still denervated and no effect of T3 on muscle reinnervation was detected at this stage of nerve repair. In contrast, after 14 weeks of nerve regeneration, T3 clearly enhanced the reinnervation of gastrocnemius and plantar EPs, demonstrated by significantly higher recovery of size and shape complexity of reinnervated EPs and also by increased acetylcholine receptor (AChRs) density on post synaptic membranes compared to PBS-treated EPs. The stimulating effect of T3 on EP reinnervation is confirmed by a higher index of compound muscle action potentials recorded in gastrocnemius muscles. In conclusion, our results provide for the first time strong evidence that T3 enhances the restoration of NMJ structure and improves synaptic transmission.

Molecular characterization of hepatitis A virus isolates from Goiânia, Goiás, Brazil

Hepatitis A virus (HAV) infection is a public health problem worldwide and the virus has been classified into six genotypes. In Brazil, the only genotype that has been found is genotype I, predominately from subgenotype IA. Here, the HAV genotypes were analyzed of 18 isolates circulating between 1996-2001 in Goiânia, state of Goiás, Brazil. Viral RNA was extracted from 18 serum samples and amplified (RT-PCR/nested-PCR), followed by the genomic sequencing of the VP1/2A junction region of the HAV genome. Sequences of 168 nucleotides were compared and analyzed using the BLAST N, Clustal X and PAUP v. 4.10b programs. All samples were classified as genotype I, with 10 belonging to subgenotype IA and eight to subgenotype IB. The subgenotype IA isolates showed greater diversity than the subgenotype IB isolates at the nucleotide level. Elevated identity values were found between isolates obtained in this study and those from other regions of the world, including Brazil, highlighting the high conservation among different isolates of this virus. However, changes in the HAV subgenotype circulation could also be observed during the evaluated period.

Eficacia de un programa de entrenamiento intradiálisis de fuerza-resistencia en combinación con electroestimulación neuromuscular: mejora en la capacidad funcional, fuerza, y calidad de vida

The purpose of this study was to determine the efficacy of a programme of strength-stamina exercises during haemodialysis, in improving muscular strength, quality of life and functional capacity to carry out everyday activities. A quantitative, experimental pre-test and post-test study was carried out. A programme of strength-stamina exercises in combination with neuromuscular electrostimulation was applied to 10 patients undergoing haemodialysis. These were three simple exercises adapted to the position in which haemodialysis was carried out. All the patients showed a significant improvement in strength, measured using functional tests to carry out everyday activities: walking (6-MWT) and sit-to-stand tests (10-STS). These tests were measured before and after the training programme. They also showed an improvement in the physical dimension of the quality of life measured using the specific questionnaire for renal patients, KDQOL-SFTM.

Application of compositional data analysis to geochemical data of marine sediments

In an earlier investigation (Burger et al., 2000) five sediment cores near the RodriguesTriple Junction in the Indian Ocean were studied applying classical statistical methods(fuzzy c-means clustering, linear mixing model, principal component analysis) for theextraction of endmembers and evaluating the spatial and temporal variation ofgeochemical signals. Three main factors of sedimentation were expected by the marinegeologists: a volcano-genetic, a hydro-hydrothermal and an ultra-basic factor. Thedisplay of fuzzy membership values and/or factor scores versus depth providedconsistent results for two factors only; the ultra-basic component could not beidentified. The reason for this may be that only traditional statistical methods wereapplied, i.e. the untransformed components were used and the cosine-theta coefficient assimilarity measure.During the last decade considerable progress in compositional data analysis was madeand many case studies were published using new tools for exploratory analysis of thesedata. Therefore it makes sense to check if the application of suitable data transformations,reduction of the D-part simplex to two or three factors and visualinterpretation of the factor scores would lead to a revision of earlier results and toanswers to open questions . In this paper we follow the lines of a paper of R. Tolosana-Delgado et al. (2005) starting with a problem-oriented interpretation of the biplotscattergram, extracting compositional factors, ilr-transformation of the components andvisualization of the factor scores in a spatial context: The compositional factors will beplotted versus depth (time) of the core samples in order to facilitate the identification ofthe expected sources of the sedimentary process.Kew words: compositional data analysis, biplot, deep sea sediments

F+0 renography in neonates and infants younger than 6 months: an accurate method to diagnose severe obstructive uropathy.

We studied the response to F+0 renography and the relative and absolute individual kidney function in neonates and < 6-mo-old infants before and after surgery for unilateral ureteropelvic junction obstruction (UJO). METHODS: The results obtained at diagnosis and after pyeloplasty for 9 children (8 boys, 1 girl; age range, 0.8-5.9 mo; mean age +/- SD, 2.4 +/- 1.5 mo) with proven unilateral UJO (i.e., affected kidney [AK]) and an unremarkable contralateral kidney (i.e., normal kidney [NK]) were evaluated and compared with a control group of 10 children (6 boys, 4 girls; age range, 0.8-2.8 mo; mean age, 1.5 +/- 0.7 mo) selected because of symmetric renal function, absence of vesicoureteral reflux or infection, and an initially dilated but not obstructed renal pelvis as proven by follow-up. Renography was performed for 20 min after injection of (123)I-hippuran (OIH) (0.5-1.0 MBq/kg) immediately followed by furosemide (1 mg/kg). The relative and absolute renal functions and the response to furosemide were measured on background-subtracted and depth-corrected renograms. The response to furosemide was quantified by an elimination index (EI), defined as the ratio of the 3- to 20-min activities: An EI > or = 3 was considered definitively normal and an EI < or = 1 definitively abnormal. If EI was equivocal (1 < EI < 3), the response to gravity-assisted drainage was used to differentiate AKs from NKs. Absolute separate renal function was measured by an accumulation index (AI), defined as the percentage of (123)I-OIH (%ID) extracted by the kidney 30-90 s after maximal cardiac activity. RESULTS: All AKs had definitively abnormal EIs at diagnosis (mean, 0.56 +/- 0.12) and were significantly lower than the EIs of the NKs (mean, 3.24 +/- 1.88) and of the 20 control kidneys (mean, 3.81 +/- 1.97; P < 0.001). The EIs of the AKs significantly improved (mean, 2.81 +/- 0.64; P < 0.05) after pyeloplasty. At diagnosis, the AIs of the AKs were significantly lower (mean, 6.31 +/- 2.33 %ID) than the AIs of the NKs (mean, 9.43 +/- 1.12 %ID) and of the control kidneys (mean, 9.05 +/- 1.17 %ID; P < 0.05). The AIs of the AKs increased at follow-up (mean, 7.81 +/- 2.23 %ID) but remained lower than those of the NKs (mean, 10.75 +/- 1.35 %ID; P < 0.05). CONCLUSION: In neonates and infants younger than 6 mo, (123)I-OIH renography with early furosemide injection (F+0) allowed us to reliably diagnose AKs and to determine if parenchymal function was normal or impaired and if it improved after surgery.

Dilatation pyélocalicielle chez le nouveau-né: attitude pratique pour le pédiatre [Antenatally detected hydronephrosis: practical approach for the pediatrician]

Approximately 1% of the fetuses present some dilatation of their urinary tract in utero. More than 50% of these antenatally detected hydronephrosis will disappear spontaneously after birth. The other 50% comprises ureteropelvic junction obstruction, vesico-ureteral reflux and primary megaureters. Postnatal radiological evaluation (renal ultrasonography and VCUG) is performed in every infant with a significantly dilated renal pelvis (> 8 mm between 20 and 30 weeks or > 10 mm after 30 weeks in utero). Renal nuclear scan should be done in every child with significant/worsening post-natal hydronephrosis. Antibioprophylaxis will be started from birth to prevent urinary tract infection. Medical or surgical approach will be chosen in the light of the uroradiological exam results and the clinical progress.

Esophageal leaks repaired by a muscle onlay approach in the presence of mediastinal sepsis.

BACKGROUND: Nineteen patients were evaluated after closure of intrathoracic esophageal leaks by a pediculated muscle flap onlay repair in the presence of mediastinal and systemic sepsis. METHODS: Intrathoracic esophageal leaks with mediastinitis and systemic sepsis occurred after delayed spontaneous perforations (n = 7) or surgical and endoscopic interventions (n = 12). Six patients presented with fulminant anastomotic leaks. Seven patients had previous attempts to close the leak by surgery (n = 4) or stenting (2) or both (n = 1). The debrided defects measured up to 2 x 12 cm or involved three quarters of the anastomotic circumference and were closed either by a full thickness diaphragmatic flap (n = 13) or a pediculated intrathoracically transposed extrathoracic muscle flap (n = 6). All patients had postoperative contrast esophagography between days 7 and 10 and an endoscopic evaluation 4 to 6 months after surgery. RESULTS: There was no 30-day mortality. During follow-up (4 to 42 months), 16 patients (84%) revealed functional and morphological restoration of the esophagointestinal integrity without further interventions. One patient required serial dilatations for a stricture, and 1 underwent temporary stenting for a persistent fistula; both patients had normal control endoscopy during follow-up. A third patient requiring permanent stenting for stenosis died from gastrointestinal bleeding due to stent erosion during follow-up. CONCLUSIONS: Intrathoracic esophageal leaks may be closed efficiently by a muscle flap onlay approach in the presence of mediastinitis and where a primary repair seems risky. The same holds true for fulminant intrathoracic anastomotic leaks after esophagectomy or other surgical interventions at the gastroesophageal junction.

Hyperglycemia downregulates Connexin36 in pancreatic islets via the upregulation of ICER-1/ICER-1γ.

Channels formed by the gap junction protein Connexin36 (CX36) contribute to the proper control of insulin secretion. We previously demonstrated that chronic exposure to glucose decreases Cx36 levels in insulin-secreting cells in vitro. Here, we investigated whether hyperglycemia also regulates Cx36 in vivo. Using a model of continuous glucose infusion in adult rats, we showed that prolonged (24-48 h) hyperglycemia reduced the Cx36 gene Gjd2 mRNA levels in pancreatic islets. Accordingly, prolonged exposure to high glucose concentrations also reduced the expression and function of Cx36 in the rat insulin-producing INS-1E cell line. The glucose effect was blocked after inhibition of the cAMP/PKA pathway and was associated with an overexpression of the inducible cAMP early repressor ICER-1/ICER-1γ, which binds to a functional cAMP-response element in the promoter of the Cx36 gene Gjd2. The involvement of this repressor was further demonstrated using an antisense strategy of ICER-1 inhibition, which prevented glucose-induced downregulation of Cx36. The data indicate that chronic exposure to glucose alters the in vivo expression of Cx36 by the insulin-producing β-cells through ICER-1/ICER-1γ overexpression. This mechanism may contribute to the reduced glucose sensitivity and altered insulin secretion, which contribute to the pathophysiology of diabetes.

NS2 Proteins of GB Virus B and Hepatitis C Virus Share Common Protease Activities and Membrane Topologies.

GB virus B (GBV-B), which is hepatotropic in experimentally infected small New World primates, is a member of the Hepacivirus genus but phylogenetically relatively distant from hepatitis C virus (HCV). To gain insights into the role and specificity of hepaciviral nonstructural protein 2 (NS2), which is required for HCV polyprotein processing and particle morphogenesis, we investigated whether NS2 structural and functional features are conserved between HCV and GBV-B. We found that GBV-B NS2, like HCV NS2, has cysteine protease activity responsible for cleavage at the NS2/NS3 junction, and we experimentally confirmed the location of this junction within the viral polyprotein. A model for GBV-B NS2 membrane topology was experimentally established by determining the membrane association properties of NS2 segments fused to green fluorescent protein (GFP) and their nuclear magnetic resonance structures using synthetic peptides as well as by applying an N-glycosylation scanning approach. Similar glycosylation studies confirmed the HCV NS2 organization. Together, our data show that despite limited amino acid sequence similarity, GBV-B and HCV NS2 proteins share a membrane topology with 3 N-terminal transmembrane segments, which is also predicted to apply to other recently discovered hepaciviruses. Based on these data and using trans-complementation systems, we found that intragenotypic hybrid NS2 proteins with heterologous N-terminal membrane segments were able to efficiently trans-complement an assembly-deficient HCV mutant with a point mutation in the NS2 C-terminal domain, while GBV-B/HCV or intergenotypic NS2 chimeras were not. These studies indicate that virus- and genotype-specific intramolecular interactions between N- and C-terminal domains of NS2 are critically involved in HCV morphogenesis. IMPORTANCE: Nonstructural protein 2 (NS2) of hepatitis C virus (HCV) is a multifunctional protein critically involved in polyprotein processing and virion morphogenesis. To gain insights into NS2 mechanisms of action, we investigated whether NS2 structural and functional features are conserved between HCV and GB virus B (GBV-B), a phylogenetically relatively distant primate hepacivirus. We showed that GBV-B NS2, like HCV NS2, carries cysteine protease activity. We experimentally established a model for GBV-B NS2 membrane topology and demonstrated that despite limited sequence similarity, GBV-B and HCV NS2 share an organization with three N-terminal transmembrane segments. We found that the role of HCV NS2 in particle assembly is genotype specific and relies on critical interactions between its N- and C-terminal domains. This first comparative analysis of NS2 proteins from two hepaciviruses and our structural predictions of NS2 from other newly identified mammal hepaciviruses highlight conserved key features of the hepaciviral life cycle.