Functional features defining the efficacy of cholesterol-conjugated, self-deliverable, chemically modified siRNAs

Progress in oligonucleotide chemistry has produced a shift in the nature of siRNA used, from formulated, minimally modified siRNAs, to unformulated, heavily modified siRNA conjugates. The introduction of extensive chemical modifications is essential for conjugate-mediated delivery. Modifications have a significant impact on siRNA efficacy through interference with recognition and processing by RNAi enzymatic machinery, severely restricting the sequence space available for siRNA design. Many algorithms available publicly can successfully predict the activity of non-modified siRNAs, but the efficiency of the algorithms for designing heavily modified siRNAs has never been systematically evaluated experimentally. Here we screened 356 cholesterol-conjugated siRNAs with extensive modifications and developed a linear regression-based algorithm that effectively predicts siRNA activity using two independent datasets. We further demonstrate that predictive determinants for modified and non-modified siRNAs differ substantially. The algorithm developed from the non-modified siRNAs dataset has no predictive power for modified siRNAs and vice versa. In the context of heavily modified siRNAs, the introduction of chemical asymmetry fully eliminates the requirement for thermodynamic bias, the major determinant for non-modified siRNA efficacy. Finally, we demonstrate that in addition to the sequence of the target site, the accessibility of the neighboring 3\u27 region significantly contributes to siRNA efficacy

Arrhythmogenic effects of hypoglycemia

Hypoglycemia is a frequent event in patients on insulin therapy. Current clinical and experimental evidence shows hypoglycemia tobe a cause of arrhythmia and, possibly, a link to increased mortality risk in patients with diabetes mellitus. This review addressesprobable mechanisms and pathogenic factors of arrhythmia development due to hypoglycemic events. We adduce data accumulatedon rates of hypoglycemia, as well as their correlation with cardiovascular and general mortality according to ACCORD, ADVANCEand VADT trials

Уровни циклического 3'- Б'-аденозинмонофосфата лимфоцитов и состояние иммунного статуса в динамике различных вариантов течения пневмонии

As a result of executed studies of the functional activity of the adenylate cyclase system (ACS) of periferal lymphocites and the immunological status condition in patients with various cources of pneumonia, the stagiality of changes of basal intracellular c-AMP levels (pmoll/106 cells), the beta-2-adrenoreception activity, and lymphocitarial response to concavaline A was revealed.The activity decrease of ACS at the beginning of pneumonia was correlated with the diminishing of such parameters as E-ROC, Ea-ROC, EAC-ROC, C3 complement fraction and with changes of levels of IgA, M, G, that is more expressed in patients with severe forms of the disease.The tendency to the activity repairment of ACS after 7—10 days of hospital treatment was noted in patients with uncomplecated entities of pneumonia accompanied by immunological status normalization. The tendency to the adenylate cyclase activity repairment was noted also in patients cured with hyperimmune antistaphylococcus serum, that was not signed in patients with the complicated disease cured with traditional treatment only.Afetr 21 days, in the group of patients with uncomplicated pneumonia cured with sustitutional immunological correction, the ACS activity coincided with immunological and clinical normalization of parameters. Firmy impairments of immunological parameters and the severity of clinical manifestation of the disease were accompanied by tyhe absence of similar c-AMP dynamics in patients with complicated pneumonia entities not cured with immunocorrection methods.The dynamics of the lymphocite ASC activity of patients with various pneumonia forms demonstrates the process of clinical immunological changes and points to the positive influence of the proper usage of immunocorrection treatment on molecular mechanisms of immunity regulation during the mentioned disease.В результате проведенных исследований функциональной активности аденилатциклазной (АЦ) системы периферических лимфоцитов и состояния иммунного статуса у пациентов с различным течением пневмонии нами выявлена этапность изменений базальных внутриклеточных уровней цАМФ (pmoll/106 cell), активности бета-2-адренорецепторов и ответа лимфоцитов на конканавалин А.Падение активности АЦ-системы в начале заболевания пневмонией коррелировало со снижением таких показателей, как Е-РОК, Еа-РОК, ЕАС-РОК, СЗ-фракции комплемента и изменениями уровней IgA, М, G, более выраженным у пациентов с тяжелыми формами заболевания.Тенденция к восстановлению активности АЦ-системы на 7—10-е сутки стационарного лечения отмечена у пациентов с неосложненными вариантами пневмонии, чему сопутствовала нормализация иммунного статуса. Отмечена тенденция к восстановлению показателей активности аденилатциклазы и у пациентов, лечившихся гипериммунной антистафилококковой плазмой, что не имело места у больных осложненными формами заболевания, в лечении которых применялись только традиционные средства.На 21-е сутки восстановление активности АЦ-системы в группе неосложненных пневмоний у пациентов, которым проводилась заместительная иммунокоррекция, совпало с нормализацией иммунологических и клинических показателей. Отсутствию подобной динамики цАМФ у пациентов с осложненными вариантами пневмонии, в лечении которых не применялись средства иммунокоррекции, сопутствовали наличие стойких нарушений иммунологических показателей и тяжесть клинических проявлений заболевания.Динамика показателей активности АЦ-системы лимфоцитов пациентов с различными вариантами пневмонии отражает процесс клинико-иммунологических изменений и указывает на положительную роль адекватного применения средств иммунокоррекции на молекулярные механизмы регуляции иммунитета при данном заболевании

Manipulation of Panx1 Activity Increases the Engraftment of Transplanted Lacrimal Gland Epithelial Progenitor Cells

Sjögren's syndrome is a systemic chronic autoimmune inflammatory disease that primarily targets the salivary and lacrimal glands (LGs). Currently there is no cure; therefore, cell-based regenerative therapy may be a viable option. LG inflammation is facilitated by extracellular ATP and mediated by the Pannexin-1 (Panx1) membrane channel glycoprotein. We propose that suppression of inflammation through manipulation of Panx1 activity can stimulate epithelial cell progenitor (EPCP) engraftment. The expression of pannexins in the mouse and human LG was assayed by qRT-PCR and immunostaining. Acute LG inflammation was induced by interleukin-1α (IL1α) injection. Prior to EPCP transplantation, IL1α-injured or chronically inflamed LGs of thrombospondin-1-null mice (TSP-1-/-) were treated with the Panx1-specific blocking peptide (10panx) or the self-deliverable RNAi (sdRNAi). The efficacy of cell engraftment and the area of inflammation were analyzed by microscopy. Panx1 and Panx2 were detected in the mouse and human LGs. Panx1 and proinflammatory factors were upregulated during acute inflammation at days 1 to 3 after the IL1α injection. The analysis of EPCP engraftment demonstrated a significant and reproducible positive correlation between the 10panx peptide or Panx1 sdRNAi treatment and the number of engrafted cells. Similarly, treatment of the LG of the TSP-1-/- mouse (mouse model of chronic LG inflammation) by either Panx1 or Caspase-4 (also known as Casp11) sdRNAi showed a significant decrease in expression of proinflammatory markers and the lymphocyte infiltration. Our results suggest that blocking Panx1 and/or Casp4 activities is a beneficial strategy to enhance donor cell engraftment and LG regeneration through the reduction of inflammation