[Resolution of the Council of Experts «Bioregulatory drugs in the complex treatment of common ENT diseases¼]. / Rezolyutsiya Soveta ekspertov «Bioregulyatsionnye preparaty v kompleksnoi terapii rasprostranennykh LOR-zabolevanii¼.

Within the framework of the III All-Russian scientific and practical conference of otorhinolaryngologists, audiologists, somnologists and maxillofacial surgeons with international participation, a meeting of experts in the field of ENT diseases was held with the support of Heel RUS LLC, October 17-18, 2019, Kazan.

SARS-CoV-2 genome analysis of strains in Pakistan reveals GH, S and L clade strains at the start of the pandemic

ObjectivesPakistan has a high infectious disease burden with about 265,000 reported cases of COVID-19. We investigated the genomic diversity of SARS-CoV-2 strains and present the first data on viruses circulating in the country. MethodsWe performed whole-genome sequencing and data analysis of SARS-CoV-2 eleven strains isolated in March and May. ResultsStrains from travelers clustered with those from China, Saudi Arabia, India, USA and Australia. Five of eight SARS-CoV-2 strains were GH clade with Spike glycoprotein D614G, Ns3 gene Q57H, and RNA dependent RNA polymerase (RdRp) P4715L mutations. Two were S (ORF8 L84S and N S202N) and three were L clade and one was an I clade strain. One GH and one L strain each displayed Orf1ab L3606F indicating further evolutionary transitions. ConclusionsThis data reveals SARS-CoV-2 strains of L, G, S and I have been circulating in Pakistan from March, at the start of the pandemic. It indicates viral diversity regarding infection in this populous region. Continuing molecular genomic surveillance of SARS-CoV-2 in the context of disease severity will be important to understand virus transmission patterns and host related determinants of COVID-19 in Pakistan.

Mutational landscape of head and neck squamous cell carcinomas in a South Asian population.

Head and neck squamous cell carcinoma (HNSCC) is the sixth most common cancer type globally and contributes significantly to burden of disease in South Asia. In Pakistan, HNSCC is among the most commonly diagnosed cancer in males and females. The increasing regional burden of HNSCC along with a unique set of risk factors merited a deeper investigation of the disease at the genomic level. Whole exome sequencing of HNSCC samples and matched normal genomic DNA analysis (n=7) was performed. Significant somatic single nucleotide variants (SNVs) were identified and pathway analysis performed to determine frequently affected signaling pathways. We identified significant, novel recurrent mutations in ASNS (asparagine synthetase) that may affect substrate binding, and variants in driver genes including TP53, PIK3CA, FGFR2, ARID2, MLL3, MYC and ALK. Using the IntOGen platform, we identified MAP kinase, cell cycle, actin cytoskeleton regulation, PI3K-Akt signaling and other pathways in cancer as affected in the samples. This data is the first of its kind from the Pakistani population. The results of this study can guide a better mechanistic understanding of HNSCC in the population, ultimately contributing new, rational therapeutic targets for the treatment of the disease.

Acute kidney injury in neonatal intensive care: Medicines involved.

BACKGROUND: The incidence of acute kidney injury (AKI) in neonates in the intensive care units and neonatal intensive care (NICU) according Plotz et al. ranges from 8% to 22% [3]. According to Andreoli, neonatal death due to AKI in NICU amounts up to 10-61% [1]. It should be in the reasons of AKI emphasize.The role of certain drugs, which are widely used in modern neonatology: nonsteroidal anti-inflammatory drugs (NSAIDs), antibiotics (aminoglycosides, glycopeptides, carbapenems, 3rd generation cephalosporins), furosemide, enalapril, in contributing to AKI should be emphasized [2]. OBJECTIVE: To identify risk factors for acute kidney injury in neonates in intensive care units and intensive care. METHODS: We performed a prospective observational case-control study of full-term newborns who were treated in the intensive care unit and neonatal intensive care of the "Children's city hospital №1" Kazan and NICU №3 "Children's Republican Clinical Hospital" in 2011-2014 years.The study included 86 term infants in critical condition, who were hospitalized to the NICU on the first days of life, - the main group. The main criterion of AKI in neonates according to neonatal AKIN classification (2011) is a serum creatinine concentration ≥1.5 mg/dL. We subdivided the main group into two subgroups:subgroup I, AKI+ consisted of 12 term infants in critical condition with the serum creatinine level ≥ 1,5 mg/dL at the age of not younger than 48 hours after birth, which was 14% of all full-term newborns who were at the NICU;subgroup II, AKI- consisted of 74 term infants in critical condition with the serum creatinine level <1.5 mg/dL at the age of not younger than 48 hours after birth.The control group was formed by random sampling, it consisted of 26 somatically healthy term infants. We used conventional methods for evaluating renal function, and enzyme immunoassay (ELISA) for the urine biomarker of AKI, IL-18.Statistical analysis was performed using SPSS, Statistics 20, and the IBM and Microsoft Office Excel 2007. The study results were subjected to statistical analysis using parametric and non-parametric methods of analysis. We present the findings as arithmetic means (M) with, standard deviation (σ) and standard error of the mean (m) according to standard formulas. RESULTS: All children were admitted to primary and emergency care with subsequent transfer to the NICU at 1-2 days of life and further treatment in the department of pathology of newborns (DPN). The duration of hospitalization of infants at the NICU for the main group averaged 5,9 ± 0,44 days; at the DPN (subsequent stage of nursing) - 11,42 ± 0,51 days; for the subgroup I, AKI+ newborns these were 7,83 ± 1,23 days and 13,75 ± 3,34 days, respectively; for the subgroup II, AKI- newborns these were 5,58 ± 0,47 (3-30) and 11,04 ± 0,3 (0-47) days, respectively. Neonates received daily average of 16,5 ± 0,3 various medicines while at NICU and 9,1 ± 0,7 while at DPN. Overall, over the entire period of hospitalization neonates of the main group received on average 25,6 ± 1,8 medicines. Of these, 2,9 ± 0,4 drugs were antibiotics, possessing nephrotoxic properties (aminoglycosides, cephalosporins, carbapenems, fluoroquinolones). Children of the main group in 100% (n = 86) of cases were treated with 3rd generation cephalosporins (ceftriaxone, cephaperazone/sulbactam (sulperazon)), in 55% od cases (n = 47) - with aminoglycosides (amikacin, gentamicin), in 1% (n = 1) - with vancomycin, in 7% (n = 6) - with carbapenems. Diuretics were prescribed to 57% (n = 49) of infants. Often, patients were treated with a combination of nephrotoxic medications. Aminoglycoside were prescribed statistically more often to neonates of the subgroup I, than of the subgroup II (p <0.01). Diuretic drugs were used more frequently and for longer periods of time in neonates of the subgroup I (AKI+), than in newborns of the subgroup II (AKI-), namely, in 83% (n = 10) for 4,6 ± 1.34 days versus 53% of cases (n = 39) for 2.84 ± 0.49 days, respectively (p <0.05). IL-18 urine level in neonates of subgroup I (AKI+) was 2 times higher than that in neonates of the subgroup II (AKI-), and 13 times higher than in neonates of the control group.The fact that the IL-18 urine level increased with progression of kidney damage, caused by nephrotoxic therapy, suggests that a significant role in the development and progression of AKI in neonates at NICU belongs to drug therapy. CONCLUSIONS: Full-term newborns in intensive care units are at high risk of AKI when they are treated with aminoglycosides in combination with diuretics for longer than 4.5 days.

Clinical conferences for physicians: Who sets the agenda?

BACKGROUND: Clinical conferences are generally defined as scheduled events at which practicing physicians themselves present to their colleagues interesting clinical cases, share their new experiences and learn about the latest achievements of medical science and practice. The value of a clinical conference is thought to be in direct communication between physicians, in analysis of topical issues in a given specialty with the aim to improve the quality of care. Speakers based on their own observations and studies reveal the most urgent problems, analyze results and offer potential decisions to their colleagues interested in the same questions. The event format may be different: workshops, highly specialized sections, round tables and seminars with participation of the leading specialists in a given field. These conferences are generally organised by the Ministries and Departments of Health, by leading research and/or educational institutions in the field, by recognised medical centres and other institutions. Recently pharmaceutical companies got actively involved in medical events, acting as sponsors of various scientific conferences and congresses, however threatening the mission of these events [1]. This brings up some uneasy questions: who are the medical conferences for? Who is in charge of setting the conference agenda? Do they contribute to evidence-based medicine; do they contribute to better health? Unfortunately, there is a trend to duplication or multiplication of conferences: various agencies and departments deliver the same conferences, presentations at which are often pre-arranged by pharmaceutical companies and do not have clear scientific novelty, while the conferences themselves have largely transformed into advertising of new pharmaceuticals or new technologies [2]. Pharmaceutical corporations sponsor invited speakers paying for their trips and paying honoraria, organising cocktail parties as part of medical activities. With the help of leading experts with impressive titles serving as speakers at the conferences, pharmaceutical companies are trying to be as close as possible to routine practice of prescribing of certain drugs, manipulating evidence, controlling scientific societies as well as the process of clinical guideline development and publication of research results [3]. The degree of expert involvement depends on their level of influence [4]. OBJECTIVE: We aimed to study how often regular medical practitioners attend these conferences; to analyse who were keynote speakers and where they were coming from; to identify which organizations were responsible for organisation of these conferences and for sending out invitations to these conferences and for disseminating information about them. METHODS: We summarized all the invitations (printed on paper) received by one regular medical practitioner employed with the outpatient clinical of the city of Kazan for the period of two years (2012-2013). RESULTS: During the study period (2012-2013), a regular medical practitioner received 47 printed paper invitations to scientific conferences: 22 in 2012 and 25 in 2013. The conferences were not distributed evenly over the months of the years. November appeared to be the month with the highest density/number of medical conferences: 7 conferences in 2012 and 10 in 2013. If the distribution was even, then we could calculate the number per month dividing the number per year by 9 active months (excluding July, August and September). This resulted in 2.4 and 2.8 conferences per month. Among these studied conferences 4 were organized by public health agencies: invitation tickets were accompanied by the corresponding official order to organise a conference, issued by the Health Department. Noteworthy, that 2 of these conferences were held in conference rooms of the largest hotels of the city. Forty-one out of 47 medical conferences were sponsored by big pharma: either jointly with the major medical higher educational institutions of the city or plainly by pharmaceutical companies. Seventeen conferences were held during official working hours, in the first half of the day. Not only the logo of the pharmaceutical companies was printed on invitation tickets, but there was also an advert of the promoted pharmaceutical brand.Nine conference invitations contained invitation to dinner. In one of the invitations to a conference on neuroscience it was written: "dinner under the unforgettable music". Two conference invitations contained invitation to a lunch. Programs of 20 conferences (which were included) listed guest lecturers, coming from the leading medical universities in Moscow and St. Petersburg. Opinion leaders' involvement: some of the leading experts acted as speakers from 4 to 7 conferences a month in this sample conference invitations package of a regular polyclinic physician. CONCLUSIONS: In 2012-2013 health practitioners were invited to attend medical conferences regularly, at least 2 times a month, with November being the busiest month. The keynote speakers were the opinion leaders from the local medical educational institutions and visitors from Moscow and St. Petersburg; their involvement with the conferences was repetitive. Governmental institutions jointly with big pharma were responsible for organisation of these conferences and attracting audience.Limitations of these observations:Unfortunately, the information on printed-paper conference invitations was not complete because not all tickets have survived. From the interview with the physician we know that in addition to these printed on paper invitations there were many invitations and alerts sent out by e-mail, SMS messages and personal phone calls, making the regularity of these conferences much higher. The physician, who kindly provided this information to us, asked not to be named or thanked in any public presentation of the results of these analyses.

[Palivizumab: four seasons in Russia].

In 2010, the Russian Federation (RF) registered palivizumab--innovative drug, based on monoclonal antibodies for passive immunization of seasonal respiratory syncytial virus (RSV) infection in children of disease severe progress risk group, which include primarily premature infants, children with bronchopulmonary dysplasia and hemodynamically significant congenital heart disease. Currently, palivizumab is included in the list of recommended medicines and medical care standards of different countries, including Russia. In the review the results of Russian research on the progress of RSV infection, its epidemiology and immunization experience gained over the 2010-2014 period are summarized in relation to the foreign data. During the four epidemic seasons palivizumab immunization covered more than 3,200 children of severe RSV infection risk group with a progressive annual increase in the number of patients who received the drug. Geography of palivizumab immunization is also greatly expanded in our country during this time. If during the first two seasons measures of immunization were taken mainly in Moscow and St. Petersburg, at the present time, thirty one territorial entities of the Russian Federation have the experience in the drug application. Analysis of the results of RSV infection immunization (made in several regions) confirms the high clinical efficacy and palivizumab safety already demonstrated in international studies. In addition, the analysis presents the potential to improve the efficiency of the integrated RSV infection immunization programs, realizing in the establishment of high-risk child group register, adequate counseling for parents, as well as the development of the routing of patients and coordination of interaction between different health institutions during the immunization.

TAK1 is required for TGF-beta 1-mediated regulation of matrix metalloproteinase-9 and metastasis.

Transforming growth factor-beta 1 (TGF-beta1) signaling in tumor cells has been implicated in tumor angiogenesis and metastasis by regulating matrix proteolysis. Although MMP-9/gelatinase-B is an important component of these TGF-beta1 responses, the mechanism of its regulation is not well understood. Here, we present evidence that TGF-beta-activated protein kinase 1 (TAK1) is critical for TGF-beta regulation of MMP-9 and the metastatic potential of breast cancer cell line MDA-MB-231. We found that suppression of TAK1 signaling by dominant-negative (dn) TAK1 or RNA interference (siRNA) reduces expression of MMP-9 and tumor cell invasion, without growth inhibition in cell culture. The orthotopic xenograft studies in SCID mice showed that suppression of TAK1 signaling by dn-TAK1 reduces tumor growth and formation of lung metastases. Dn-TAK1 reduced the proliferation Ki-67 index and neovasculature of orthotopic xenografts. TAK1-mediated regulation of MMP-9 involves NF-kappaB signaling. Dn-TAK1 reduces NF-kappaB transcriptional response and inhibition of NF-kappaB reduces expression of MMP-9 and activity of the MMP-9 promoter reporter. Together, these findings suggest that TAK1 contributes to TGF-beta1-mediated tumor angiogenesis and metastasis via a mechanism involving the TAK1-NF-kappaB-MMP-9 pathway.

ALK5 promotes tumor angiogenesis by upregulating matrix metalloproteinase-9 in tumor cells.

Transforming growth factor beta 1 (TGF-beta1) is a potent tumor suppressor but, paradoxically, TGF-beta1 enhances tumor growth and metastasis in the late stages of cancer progression. This study investigated the role of TGF-beta type I receptor, ALK5, and three mitogen-activated protein kinases (MAPKs) in metastasis by breast cancer cell line MDA-MB-231. We show that autocrine TGF-beta signaling in MDA-MB-231 cells is required for tumor cell invasion and tumor angiogenesis. Expression of kinase-inactive ALK5 reduces tumor invasion and formation of new blood vessels within the tumor orthotopic xenografts in severe combined immunodeficiency (SCID) mice. In contrast, constitutively active ALK5-T204D enhances tumor invasion and angiogenesis by stimulating expression of matrix metalloproteinase MMP-9/gelatinase-B. Ablation of MMP-9 in ALK5-T204D cells by RNA interference (RNAi) reduces tumor invasion and tumor growth. Importantly, RNAi-MMP-9 reduces tumor neovasculature and increases tumor cell death. Induction of MMP-9 by TGF-beta-ALK5 signaling requires MEK-ERK but not JNK, p38 MAPK or Smad4. Dominant-negative MEK blocks and constitutively active MEK1 enhances MMP-9 expression. However, all three MAPK cascades (ERK, JNK and p38 MAPK) are required for TGF-beta-mediated cell migration. Collectively, our results show that TGF-beta-ALK5-MAPK signaling in tumor cells promotes tumor angiogenesis and MMP-9 is an important component of this program.

Basal induction of cytochrome P-450 1A1/1A2 in cells of the liver, small intestinal villi, and lymph nodes in rats.

Rat liver, small intestinal wall, mesenteric, mediastinal, and popliteal lymph nodes of rats were studied by indirect immunoperoxidase method with monoclonal antibodies to cytochrome P-450 1A1/1A2. These cytochrome forms were detected in low concentrations in hepatocytes, macrophages of the small intestinal villi and mesenteric and mediastinal lymph nodes of intact animals. Basal induction of monooxygenase enzymes can be caused by cytochrome inductors entering with air (environmental pollution) or food.

The possibility of cytochrome P450 1A1/1A2 induction in cells of distant lymph nodes of rats after enteral treatment with benzo[a]pyrene.

The mesenteric, mediastinal, and popliteal lymph nodes of rats were studied by indirect immunoperoxidase method using monoclonal antibodies to cytochrome P450 1A1/1A2 after oral treatment with benzo[a]pyrene. These cytochrome forms were detected in monocytes, macrophages, reticular and littoral cells, cell detritus, and liquid contents of the paracortical and medullary sinuses of all studied lymph nodes. The results indicate that exo- and endogenous toxic substances are oxidized not only in the liver, but also in lymph nodes.

[Influence of the activation of the immune system cells on the parameters of lipid metabolism in macrophages]. / Vliianie aktivatsii kletok immunnoi sistemy na parametry lipidnogo obmena v makrofagakh.

The influence of tumor necrosis factor a (TNF-alpha) and media, conditioned by activated macrophages and lymphocytes and containing a complex of biologically active compounds (including cytokines), on the parameters of lipid metabolism in macrophages was studied. The addition of recombinant TNF-alpha and immunocompetent cell-conditioned media to mouse peritoneal macrophages culture stimulated labelled oleate incorporation into cholesterol esters and triglycerides, as well as labelled glycerine incorporation into cholesterol esters, but inhibited labelled cholesterol incorporation into cholesterol esters. One of the mechanisms of the influence of activated immunocompetent cells on cholesterol metabolism in macrophages was, supposedly, the stimulation of sphigmomyelinase activity by a complex of anti-inflammatory cytokines produced by these cells on their activation.

Benzo[alpyrene induction of cytochrome P450 1A1/1A2 in the lymph nodes of rats.

Studies of mesenteric lymph nodes of rats by indirect immunoperoxidase method using monoclonal antibodies to cytochrome P450 1A/1A2 after oral dose of benzo[a]pyrene showed the presence of these cytochrome forms in monocytes, macrophages, reticular and litoral cells, cell detritus, and liquid contents of the paracortical zone and medullary substance sinuses. Oxidation of various exo- and endogenous toxins in the lymph nodes was revealed.

Bone mineralization in senescence-accelerated OXYS rats.

We compared age-related changes in the mineral composition of bone tissues in Wistar and senescence-accelerated OXYS rats. The mass concentrations of calcium, phosphorus, and magnesium were measured by atomic emission spectrometry. OXYS rats are characterized by early mineralization of the bone tissue, which peaked by the 6th month of life. Mineralization defects in 12-month-old OXYS rats included accumulation of iron and phosphorus, decrease in the Ca/P ratio, and increase in ash weight. These changes reflected early development and accelerated aging of these animals. Studies of the bone tissue in OXYS rats indicate that these animals can be used for evaluation of the mechanisms of involutional osteoporosis.

[Effect of stimulation and inhibition of macrophage functions on hypercholesterolemia in rats]. / Vliianie stimuliatsii i ingibirovaniia funktsii makrofagov na razvitie giperkholesterinemii u krys.

In this study we investigated the effects of zymosan and prodigiozan, the macrophage stimulators, and GdCl3, a macrophage inhibitor, on blood lipoprotein composition, activities of liver cholesterly ester (CE) metabolising enzymes, incorporation of [14C]cholesterol (C) into bile acids and accumulation and synthesis of CE in peritoneal macrophages (PM) of rats fed with C-enriched diet for 7 days. The increase of number of phagocyte cells quantity in liver and blood colony-stimulating activity in rats pretreated with intravenous injection of zymosan and prodigiozan was accompanied by reduced C content in blood low density and very low density lipoproteins (LDL and VLDL), increase of liver lysosomal CE hydrolase activity (without change of acyl-CoA:C acyltransferase and cytoplasmatic CE hydrolase activities) and incorporation of labeled C into bile acids and decrease of CE formation and accumulation in PM in rats with hypercholesterolemia. In contrast, reduction of phagocyte population in liver caused by intravenous injection of GdCl3 was accompanied by enhancement of C and CE level in blood LDL and VLDL and decrease of lysosomal CE hydrolase activity and incorporation of C into bile acids in liver of C-feeding rats. The data obtained suggest that the stimulation of mononuclear phagocyte system may lead to a decrease of plasma C via activation of LDL and VLDL catabolism and induction of bile acid synthesis in liver.

Carboxymethylated beta-1,3-glucan inhibits the binding and degradation of acetylated low density lipoproteins in macrophages in vitro and modulates their plasma clearance in vivo.

In atherosclerotic lesions, macrophages are transformed into foam cells accumulating modified low density lipoproteins (LDL) via the scavenger receptor pathway. We have investigated the effects of carboxymethylated beta-1,3-glucan (CMG) on acetylated LDL (AcLDL) metabolism in murine peritoneal macrophages in vitro and upon the clearance of AcLDL by rat liver in vivo. In cultured murine peritoneal macrophages, CMG reduced substantially the AcLDL-induced synthesis of cholesteryl esters, decreased the binding and degradation of [125I]-AcLDL in a dose-dependent manner with complete inhibition at 20-30 nM, but had no effect on the binding and degradation of native [125I]-LDL. In contrast, other polysaccharides studied, namely zymosan lipopolysaccharide, non-modified glucan and mannan Rhodexman, had a slight effect at concentrations significantly exceeding the concentrations of CMG. [125I]-AcLDL injected intravenously into rats was cleared from the blood with a half-life of 3.7 min. About 56 per cent of the label of injected [125I]-AcLDL was recovered in the liver 15 min after administration. Co-injection of the labelled AcLDL with CMG (25 mg kg-1 b.w.) decreased the rate of AcLDL clearance so that the half-life increased to 6.0 min. Injections of CMG (25 mg kg-1 b.w.) 48 and 24 h before the determination increased the rate of [125I]-AcLDL clearance (with a half-life of about 2.3 min) and increased the uptake of AcLDL by the liver. We suggest that CMG competed with AcLDL for scavenger receptors in vitro and in vivo and repeated CMG injections before the measurements of AcLDL resulted in the induction of scavenger receptor function.

[Endocytosis by liver cells during stimulation of the mononuclear phagocyte system by yeast polysaccharides]. / Endotsitoz kletkami pecheni pri stimuliatsii sistemy mononuklearnykh fagotsitov drozhzhevymi polisakharidami.

Endocytosis (phagocytosis, fluid-phase- and receptor-mediated endocytosis) by liver cells, lysosomal enzyme activities have been studied during macrophages stimulation by yeast polysaccharides. It was shown that like macrophages stimulator zymosan, yeast polysaccharides cryelan and rhodexman increased the carbon particles phagocytosis. The most effective was intravenous administration of yeast polysaccharides. Compared to rhodexman, the effect of cryelan was more prominent. Macrophages stimulation was followed by suppression of fluid-phase endocytosis by liver cells. Increased activity of cathepsin B was discovered on day 5 after macrophages stimulation (proteinase, most typical for macrophages enzymes).

Endocytosis by liver cells during suppression of intralysosomal proteolysis.

The lysosomotropic agent chloroquine is widely used as a specific inhibitor of intralysosomal proteolysis in isolated hepatocytes. It was shown that in vitro chloroquine reversibly inhibited purified cathepsins H, B, L in concentrations less than those observed inside lysosomes in vivo. However, administration of high doses of chloroquine to rats (30-50 mg/kg i.p. as a single or repeated injections) was followed by increased cathepsin D and cysteine proteinase activities, as well as other lysosomal enzymes. Chloroquine administration did not induce any changes of carbon particles phagocytosis by liver cells (macrophages); modifications of fluid-phase (125I-PVP uptake) and receptor-mediated endocytosis (125I-asialo-fetuin uptake) were noted. Chloroquine administered in vivo reproduced some symptoms of lysosomal storage diseases (especially during repeated drug administration).