Hepatocyte polarization is essential for the productive entry of the hepatitis B virus

Human hepatitis B virus (HBV) is characterized by a high species specificity and a distinct liver tropism. Within the liver, HBV replication occurs in differentiated and polarized hepatocytes. Accordingly, the in vitro HBV infection of primary human hepatocytes (PHHs) and the human hepatoma cell line, HepaRG, is restricted to differentiated, hepatocyte-like cells. Though preparations of PHH contain up to 100% hepatic cells, cultures of differentiated HepaRG cells are a mixture of hepatocyte-like and biliary-like epithelial cells. We used PHH and HepaRG cells and compared the influence of virus inoculation dose, cell differentiation, and polarization on productive HBV infection. At multiplicities of genome equivalents (mge) >8,000, almost 100% of PHHs could be infected. In contrast, only a subset of HepaRG cells stained positive for HBcAg at comparable or even higher mge. Infection predominantly occurred at the edges of islands of hepatocyte-like HepaRG cells. This indicates a limited accessibility of the HBV receptor, possibly as a result of its polar sorting. Multidrug resistance protein 2 (MRP2), a marker selectively transported to the apical (i.e., canalicular) cell membrane, revealed two polarization phenotypes of HepaRG cells. HBV infection within the islands of hepatocyte-like HepaRG cells preferentially occurred in cells that resemble PHH, exhibiting canalicular structures. However, disruption of cell-cell junctions allowed the additional infection of cells that do not display a PHH-like polarization. CONCLUSION: HBV enters hepatocytes via the basolateral membrane. This model, at least partially, explains the difference of PHH and HepaRG cells in infection efficacy, provides insights into natural HBV infection, and establishes a basis for optimization of the HepaRG infection system.     

Clinical relevance of RET variants G691S, L769L, S836S and S904S to sporadic medullary thyroid cancer

Background Based on reports of higher frequencies among patients with sporadic medullary thyroid cancer (MTC) relative to external controls, the RET (REarranged during Transfection) variants G691S, L767L, S836S and S904S have been considered disease modifiers, suggesting greater lifetime risks of MTC. Other studies, employing different external controls, failed to confirm this association. Using a complementary approach, this study aimed at exploring differences in clinico‐pathological characteristics among patients with sporadic MTC carrying no (wildtype), one (heterozygotes) or both (homozygotes) homologue RET variants in the germline, with wildtype cases acting as internal controls. Methods Included in this investigation were 150 patients with complete genetic information on G691S, L769L, S836S and S904S RET alleles operated on for sporadic MTC at a tertiary referral centre. Results Not one statistically significant dose–response relationship was identified between any RET variant (wildtype vs RET heterozygotes vs homologue RET homozygotes) and patient age at MTC diagnosis, gender, primary tumour size, extrathyroidal extension, numbers of involved and removed lymph nodes, or distant metastasis. L769L and S836S homozygotes, unlike G691S and S904S homozygotes, were either rare or absent, limiting the analyses to comparisons of heterozygosity versus wildtype. On time‐to‐event analysis, G691S, L769L, S836S or S904S carriers and noncarriers developed MTC at similar rates. Conclusions In carriers and noncarriers of the RET variants G691S, L767L, S836S and S904S, sporadic MTC appeared clinically and pathologically indistinguishable. This observation, along with the inconclusive evidence of previous association studies, calls for larger longitudinal association studies with age‐ and sex‐matched external controls and additional functional studies of RET biology.     

Th17 and non-Th17 interleukin-17-expressing cells in chronic lymphocytic leukemia: delineation, distribution, and clinical relevance

Background

The levels and clinical relevance of Th17 cells and other interleukin-17-producing cells have not been analyzed in chronic lymphocytic leukemia. The objective of this study was to quantify blood and tissue levels of Th17 and other interleukin-17-producing cells in patients with this disease and correlate blood levels with clinical outcome.

Design and Methods

Intracellular interleukin-17A was assessed in blood and splenic mononuclear cells from patients with chronic lymphocytic leukemia and healthy subjects using flow cytometry. Interleukin-17A-producing cells were analyzed in formalin-fixed, paraffin-embedded spleen and lymph node sections using immunohistochemistry and immunofluorescence.

Results

The absolute numbers of Th17 cells in peripheral blood mononuclear cells and the percentages of Th17 cells in spleen cell suspensions were higher in patients with chronic lymphocytic leukemia than in healthy subjects; in six out of eight paired chronic lymphocytic leukemia blood and spleen sample comparisons, Th17 cells were enriched in spleen suspensions. Circulating Th17 levels correlated with better prognostic markers and longer overall survival of the patients. Two “non-Th17” interleukin-17-expressing cells were identified in chronic lymphocytic leukemia spleens: proliferating cells of the granulocytic lineage and mature mast cells. Granulocytes and mast cells in normal spleens did not express interleukin-17. Conversely, both chronic lymphocytic leukemia and healthy lymph nodes contained similar numbers of interleukin-17+ mast cells as well as Th17 cells.

Conclusions

Th17 cells are elevated in chronic lymphocytic leukemia patients with better prognostic markers and correlate with longer survival. Furthermore, non-Th17 interleukin-17A-expressing cells exist in chronic lymphocytic leukemia spleens as maturing granulocytes and mature mast cells, suggesting that the microenvironmental milieu in leukemic spleens promotes the recruitment and/or expansion of Th17 and other IL-17-expressing cells. The pathophysiology of Th17 and non-Th17-interleukin-producing cells in chronic lymphocytic leukemia and their distributions and roles in this disease merit further study.     

The relation between lateralisation,early start of training,and amount of practice in musicians: A contribution to the problem of handedness classification

This study investigates the influence of extensive bimanual training in professional musicians on the incidence of handedness in the most basic form of right-handedness (RH) and non-right-handedness (NRH), according to Annett's “right shift theory”. The lateralisation coefficients (LCs) of a total sample of 128 bimanually performing music students were calculated for speed, regularity, and fatigue of tapping by using the speed tapping paradigm. Additionally, the accumulated amount of practice was recorded by means of retrospective interviews. The proportion of designated right-handers (dRH) and non-right-handers (dNRH) in hand performance was identified by binary logistic regression from LCs. A proportion of 30.8% designated NRH in the group of musicians was found, while in the control group of non-musicians (matched for age range) a proportion of 21.7% designated NRH was observed. Incidence of dNRH was higher in string players (35.6%) than in pianists (27.1%). As an effect of the extensive training of the left hand, tapping regularity increased and tapping fatigue decreased among those participants who evidenced an increased amount of accumulated practice time on the instrument. However, speed difference between hands (as indicated by LCs) remained uninfluenced by bimanual training. This finding is in contrast to those of Jäncke, Schlaug, and Steinmetz (1997). Finally, our study provides a more reliable (statistical) classification as an external criterion for future genetic analyses of handedness.     

Odor Identification and Self-reported Olfactory Functioning in Patients with Subtypes of Mild Cognitive Impairment

Olfactory dysfunction is a very early symptom of Alzheimer's disease (AD), and olfactory dysfunction has also been found in mild cognitive impairment (MCI). The goal of the present study was to compare odor identification ability and self-reported olfactory functioning in patients with different types of MCI. We included 104 elderly participants classified into two groups: patients with mild cognitive impairment (MCI) and elderly controls (EC). Based on their performance in neuropsychological testing the study population was divided into four groups of participants based on cognitive features: amnestic MCI single domain (11), amnestic MCI multiple domain (19), non-amnestic MCI single domain (21) and non-amnestic MCI multiple domain (13), respectively. The MCI patients were compared to 40 elderly controls (EC) controls with no cognitive deficit. Comparison for odor identification revealed a significant difference between amnestic MCI multiple domain patients and the EC group. No other group comparison was significant. Statistical analyses for self-reported olfactory functioning revealed no significant group differences between any subgroup of MCI patients and the control group. Correlational analyses indicated that odor identification ability was related to cognition whereas no relationship was found for self-reported olfactory functioning. The present study showed that amnestic MCI patients with additional deficits in other cognitive domains have a specific odor identification impairment. Together with cognitive testing, olfactory testing may more accurately help predict whether or not a patient with MCI will convert to AD in the near future.