Kindlin-3 deficiency leads to impaired erythropoiesis and erythrocyte cytoskeleton.

Kindlin-3 (K3) is critical for the activation of integrin adhesion receptors in hematopoietic cells. In humans and mice, K3 deficiency is associated with impaired immunity and bone development, bleeding, and aberrant erythrocyte shape. To delineate how K3 deficiency (K3KO) contributes to anemia and misshaped erythrocytes, mice deficient in erythroid (K3KO∖EpoR-cre) or myeloid cell K3 (K3KO∖Lyz2cre), knockin mice expressing mutant K3 (Q597W598 to AA) with reduced integrin-activation function (K3KI), and control wild-type (WT) K3 mice were studied. Both K3-deficient strains and K3KI mice showed anemia at baseline, reduced response to erythropoietin stimulation, and compromised recovery after phenylhydrazine (PHZ)-induced hemolytic anemia as compared with K3WT. Erythroid K3KO and K3 (Q597W598 to AA) showed arrested erythroid differentiation at proerythroblast stage, whereas macrophage K3KO showed decreased erythroblast numbers at all developmental stages of terminal erythroid differentiation because of reduced erythroblastic island (EBI) formation attributable to decreased expression and activation of erythroblast integrin α4ß1 and macrophage αVß3. Peripheral blood smears of K3KO∖EpoR-cre mice, but not of the other mouse strains, showed numerous aberrant tear drop-shaped erythrocytes. K3 deficiency in these erythrocytes led to disorganized actin cytoskeleton, reduced deformability, and increased osmotic fragility. Mechanistically, K3 directly interacted with F-actin through an actin-binding site K3-LK48. Taken together, these findings document that erythroid and macrophage K3 are critical contributors to erythropoiesis in an integrin-dependent manner, whereas F-actin binding to K3 maintains the membrane cytoskeletal integrity and erythrocyte biconcave shape. The dual function of K3 in erythrocytes and in EBIs establish an important functional role for K3 in normal erythroid function.

The Kindlin2-p53-SerpinB2 signaling axis is required for cellular senescence in breast cancer.

In cancer, cellular senescence is a complex process that leads to inhibition of proliferation of cells that may develop a neoplastic phenotype. A plethora of signaling pathways, when dysregulated, have been shown to elicit a senescence response. Two well-known tumor suppressor pathways, controlled by the p53 and retinoblastoma proteins, have been implicated in maintaining the cellular senescence phenotype. Kindlin-2, a member of an actin cytoskeleton organizing and integrin activator proteins, has been shown to play a key role in the regulation of several hallmarks of several cancers, including breast cancer (BC). The molecular mechanisms whereby Kindlin-2 regulates cellular senescence in BC tumors remains largely unknown. Here we show that Kindlin-2 regulates cellular senescence in part through its interaction with p53, whereby it regulates the expression of the p53-responsive genes; i.e., SerpinB2 and p21, during the induction of senescence. Our data show that knockout of Kindlin-2 via CRISPR/Cas9 in several BC cell lines significantly increases expression levels of both SerpinB2 and p21 resulting in the activation of hallmarks of cellular senescence. Mechanistically, interaction between Kindlin-2 and p53 at the promotor level is critical for the regulated expression of SerpinB2 and p21. These findings identify a previously unknown Kindlin-2/p53/SerpinB2 signaling axis that regulates cellular senescence and intervention in this axis may serve as a new therapeutic window for BCs treatment.

The Kindlin-2 regulation of epithelial-to-mesenchymal transition in breast cancer metastasis is mediated through miR-200b.

Metastasis is the main cause of death in cancer patients, including breast cancer (BC). Despite recent progress in understanding the biological and molecular determinants of BC metastasis, effective therapeutic treatments are yet to be developed. Among the multitude of molecular mechanisms that regulate cancer metastasis, the epithelial-to-mesenchymal transition (EMT) program plays a key role in the activation of the biological steps leading to the metastatic phenotype. Kindlin-2 has been associated with the pathogenesis of several types of cancers, including BC. The role of Kindlin-2 in the regulation of BC metastasis, and to a lesser extent in EMT is not well understood. In this study, we show that Kindlin-2 is closely associated with the development of the metastatic phenotype in BC. We report that knockout of Kindlin-2 in either human or mouse BC cells, significantly inhibits metastasis in both human and mouse models of BC metastasis. We also report that the Kindlin-2-mediated inhibition of metastasis is the result of inhibition of expression of key molecular markers of the EMT program. Mechanistically, we show that miR-200b, a master regulator of EMT, directly targets and inhibits the expression of Kindlin-2, leading to the subsequent inhibition of EMT and metastasis. Together, our data support the targeting of Kindlin-2 as a therapeutic strategy against BC metastasis.

αMß2 Is Antiatherogenic in Female but Not Male Mice.

Atherosclerosis is a complex inflammatory process characterized by monocyte recruitment into the arterial wall, their differentiation into macrophages, and lipid accumulation. Because integrin αMß2 (CD11b/CD18) mediates multiple diverse functions of leukocytes, we examined its role in atherogenesis. αM-/-/ApoE-/- and ApoE-/- mice were fed a control or high fat diet for 3 or 16 wk to induce atherogenesis. Unexpectedly, αM deficiency accelerated development of atherosclerosis in female but not in male mice. The size of aortic root lesions was 3-4.5-fold larger in female αM-/-/ApoE-/- than in ApoE-/- mice. Monocyte and macrophage content within the lesions was increased 2.5-fold in female αM-/-/ApoE-/- mice due to enhanced proliferation. αMß2 elimination promoted gender-dependent foam cell formation due to enhanced uptake of cholesterol by αM-/-/ApoE-/- macrophages. This difference was attributed to enhanced expression of lipid uptake receptors, CD36 and scavenger receptor A1 (SR-A1), in female mice. Macrophages from female αM-/-/ApoE-/- mice showed dramatically reduced expression of FoxM1 transcription factor and estrogen receptors (ER) α and ß. As their antagonists inhibited the effect of 17ß-estradiol (E2), E2 decreased CD36, SR-A1, and foam cell formation in ApoE-/- macrophages in an ERα- and ERß-dependent manner. However, female αM-/-/ApoE-/- macrophages failed to respond to E2 and maintained elevated CD36, SR-A1, and lipid accumulation. FoxM1 inhibition in ApoE-/- macrophages reduced ERs and enhanced CD36 and SR-A1 expression, whereas FoxM1 overexpression in αM-/-/ApoE-/- macrophages reversed their proatherogenic phenotype. We demonstrate a new, surprising atheroprotective role of αMß2 in female ApoE-/- mice. αMß2 maintains ER expression in macrophages and E2-dependent inhibition of foam cell formation.

Kindlin-2 interacts with endothelial adherens junctions to support vascular barrier integrity.

KEY POINTS: A reduction in Kindlin-2 levels in endothelial cells compromises vascular barrier function. Kindlin-2 is a previously unrecognized component of endothelial adherens junctions. By interacting directly and simultaneously with ß- or γ-catenin and cortical actin filaments, Kindlin-2 stabilizes adherens junctions. The Kindlin-2 binding sites for ß- and γ-catenin reside within its F1 and F3 subdomains. Although Kindlin-2 does not associate directly with tight junctions, its downregulation also destabilizes these junctions. Thus, impairment of both adherens and tight junctions may contribute to enhanced leakiness of vasculature in Kindlin-2+/- mice. ABSTRACT: Endothelial cells (EC) establish a physical barrier between the blood and surrounding tissue. Impairment of this barrier can occur during inflammation, ischaemia or sepsis and cause severe organ dysfunction. Kindlin-2, which is primarily recognized as a focal adhesion protein in EC, was not anticipated to have a role in vascular barrier. We tested the role of Kindlin-2 in regulating vascular integrity using several different approaches to decrease Kindlin-2 levels in EC. Reduced levels of Kindlin-2 in Kindlin-2+/- mice aortic endothelial cells (MAECs) from these mice, and human umbilical ECs (HUVEC) treated with Kindlin-2 siRNA showed enhanced basal and platelet-activating factor (PAF) or lipopolysaccharide-stimulated vascular leakage compared to wild-type (WT) counterparts. PAF preferentially disrupted the Kindlin-2+/- MAECs barrier to BSA and dextran and reduced transendothelial resistance compared to WT cells. Kindlin-2 co-localized and co-immunoprecipitated with vascular endothelial cadherin-based complexes, including ß- and γ-catenin and actin, components of adherens junctions (AJ). Direct interaction of Kindlin-2 with ß- and γ-catenin and actin was demonstrated in co-immunoprecipitation and surface plasmon resonance experiments. In thrombin-stimulated HUVECs, Kindlin-2 and cortical actin dissociated from stable AJs and redistributed to radial actin stress fibres of remodelling focal AJs. The ß- and γ-catenin binding site resides within the F1 and F3 subdomains of Kindlin-2 but not the integrin binding site in F3. These results establish a previously unrecognized and vital role of Kindlin-2 with respect to maintaining the vascular barrier by linking Vascuar endothelial cadherin-based complexes to cortical actin and thereby stabilizing AJ.

Kindlin-2 Regulates the Growth of Breast Cancer Tumors by Activating CSF-1-Mediated Macrophage Infiltration.

Interplay between tumor cells and host cells in the tumor microenvironment dictates the development of all cancers. In breast cancer, malignant cells educate host macrophages to adopt a protumorigenic phenotype. In this study, we show how the integrin-regulatory protein kindlin-2 (FERMT2) promotes metastatic progression of breast cancer through the recruitment and subversion of host macrophages. Kindlin-2 expression was elevated in breast cancer biopsy tissues where its levels correlated with reduced patient survival. On the basis of these observations, we used CRISPR/Cas9 technology to ablate Kindlin-2 expression in human MDA-MB-231 and murine 4T1 breast cancer cells. Kindlin-2 deficiency inhibited invasive and migratory properties in vitro without affecting proliferation rates. However, in vivo tumor outgrowth was inhibited by >80% in a manner associated with reduced macrophage infiltration and secretion of the macrophage attractant and growth factor colony-stimulating factor-1 (CSF-1). The observed loss of CSF-1 appeared to be caused by a more proximal deficiency in TGFß-dependent signaling in Kindlin-2-deficient cells. Collectively, our results illuminate a Kindlin-2/TGFß/CSF-1 signaling axis employed by breast cancer cells to capture host macrophage functions that drive tumor progression. Cancer Res; 77(18); 5129-41. ©2017 AACR.

[Brain-derived neurotrophic factor (BDNF) and nerve growth factor (NGF) blood levels in patients with acute carbon monoxide poisoning - a preliminary observations].

Background: Neurotrophins are the family of proteins which stimulate and regulate the process of neurogenesis. Several factors belong to the family, mainly nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT 3), and neurotrophin-4/5 (NT-4/5). Acute poisoning with carbon monoxide (CO), which usually is accompanied by neurologic symptoms, can potentially change the secretion profile of neurotrophins. Aim of the study. The main goal of the study is to assess the changes of NGF and BDNF plasma levels during an acute phase of CO poisoning as well as immediately after recovery. Additionally, the relationship among neurotrophin levels and selected aspects of clinical course of CO poisoning were studied. Materials and Methods: The study group consisted of 18 patients (mean age: 31.8±10.3 years) hospitalized in Toxicology Department of University Hospital in Cracow because of acute CO poisoning. There were 10 women (mean age: 30.2±6.9 years) and 8 men (mean age 33.9±13.7 years) in the group. The levels of NGF and BDNF were evaluated using immunoenzymatic method (ELISA) in plasma samples taken thrice in each patient. The sample 1. was taken during hospital admission, the sample 2. about 12-36 hours after admission, and the sample 3. just before the hospital discharging (usually, on the 3rd-4th day). The clinical data were collected from patients' anamnesis, physical examination and neuropsychological evaluation. The statistical analysis were performed using tools comprised in STATISTICA 12.0 PL (StatSoft Polska, Cracow, Poland) software. Results: The majority of NGF plasma levels were less than 14 pg/mL (values below the limit of quantification), contrary to the sole case of 34.3 pg/mL. BDNF plasma levels ranged from 4.8 ng/mL to above 48 ng/mL, i.e. they were higher than the upper limit of measurement range for the plasma dilution which had been used. The comparison of NGF and BDNF plasma levels in the study group with their analogues in healthy volunteers taken from the literature indicates that NGF level declines and BDNF level rises in patients with CO poisoning. The profile of BDNF concentrations in the majority of patients formed the characteristic pattern: BDNF sample 1. > BDNF sample 2. < BDNF sample 3. Taking all the values of BDNF higher than 48 ng/mL as equal to 48 ng/ mL, the statistically significant difference among 3 sample series was found according to BDNF levels. Maintaining the above mentioned assumption, the statistically significant negative correlation between the number of higher cognitive functions disturbed in one patient at the same time and the BDNF levels in sample series 2 was discovered, as well as the weak correlations between BDNF level in sample series 1 and carboxyhaemoglobin or lactate level. Moreover, weak but statistically significant correlations were present between the duration of CO exposure and BDNF levels in each sample series. Conclusions: The NGF plasma level is probably declined, while the BDNF plasma level is increased in patients with acute CO poisoning. The concentration­time curve for the plasma BDNF may sometimes undergo fluctuations with two peaks on its course. Plasma BDNF level may serve as a biological marker of disturbed higher cognitive functions in acute CO poisoning. Some clinical aspects of CO poisoning (duration of exposure, HbCO and lactate blood levels) may influence BDNF level.

[Structure of poison xenobiotic based on results of toxicological analysis from Toxicological Laboratory in Krakow in years 2003 and 2013]. / Struktura zatruc ksenobiotykami w oparciu o oznaczenia toksykologiczne wykonane w Pracowni Toksykologii w Krakowie w latach 2003 i 2013.

The aim of the study was to present changes in structure of toxicological analysis performed for hospitals in south-east Poland in last decennium. Material for the study were data from Toxicological Laboratory in Krakow in years 2003 and 2013. Total number of toxicological analysis decreased by about 30% in the period. The decline was related to analysis of ethanol, drugs of abuse (amphetamines, opiates, THC) and medicines. The number of determinations of acetylocholinoesterase activity (AChE)--marker of exposition to organophosphate and carbamate pesticides and carboxyhemoglobin (COHb)--marker of exposition to carbon monoxide also declined. The number of analysis of toxic alcohols (methanol and ethylene glycol) raised. The demand for determinations of new toxic substances (new drugs of abuse, "spices") appeared in the studied period. Advanced analytical methods (gas chromatography (GC), high performance liquid chromatography (HPLC), atomic absorption spectrometry (AAS) came into use in day-to-day routine. Changes in structure and numbers of determinations performed in regional toxicology laboratory reflects different trends. Prevalence of rapid tests for determination of drugs of abuse and alcohol in general medical laboratories caused decrease of number of the analysis in specialized toxicology laboratory. On the other hand growing demand for specialized analysis: new drugs of abuse, spices, new drugs, toxic alcohols and heavy metals was observed in last decennium.

Dual role of the leukocyte integrin αMß2 in angiogenesis.

Polymorphonuclear neutrophils (PMNs) and macrophages are crucial contributors to neovascularization, serving as a source of chemokines, growth factors, and proteases. α(M)ß(2)(CD11b/CD18) and α(L)ß(2)(CD11a/CD18) are expressed prominently and have been implicated in various responses of these cell types. Thus, we investigated the role of these ß2 integrins in angiogenesis. Angiogenesis was analyzed in wild-type (WT), α(M)-knockout (α(M)(-/-)), and α(L)-deficient (α(L)(-/-)) mice using B16F10 melanoma, RM1 prostate cancer, and Matrigel implants. In all models, vascular area was decreased by 50-70% in α(M)(-/-) mice, resulting in stunted tumor growth as compared with WT mice. In contrast, α(L) deficiency did not impair angiogenesis and tumor growth. The neovessels in α(M)(-/-) mice were leaky and immature because they lacked smooth muscle cell and pericytes. Defective angiogenesis in the α(M)(-/-) mice was associated with attenuated PMN and macrophage recruitment into tumors. In contrast to WT or the α(L)(-/-) leukocytes, the α(M)(-/-) myeloid cells showed impaired plasmin (Plm)-dependent extracellular matrix invasion, resulting from 50-75% decrease in plasminogen (Plg) binding and pericellular Plm activity. Surface plasmon resonance verified direct interaction of the α(M)I-domain, the major ligand binding site in the ß(2) integrins, with Plg. However, the α(L)I-domain failed to bind Plg. In addition, endothelial cells failed to form tubes in the presence of conditioned medium collected from TNF-α-stimulated PMNs derived from the α(M)(-/-) mice because of severely impaired degranulation and secretion of VEGF. Thus, α(M)ß(2) plays a dual role in angiogenesis, supporting not only Plm-dependent recruitment of myeloid cells to angiogenic niches, but also secretion of VEGF by these cells.

Kindlin-3 enhances breast cancer progression and metastasis by activating Twist-mediated angiogenesis.

The FERM domain containing protein Kindlin-3 has been recognized as a major regulator of integrin function in hematopoietic cells, but its role in neoplasia is totally unknown. We have examined the relationship between Kindlin-3 and breast cancer in mouse models and human tissues. Human breast tumors showed a ∼7-fold elevation in Kindlin-3 mRNA compared with nonneoplastic tissue by quantitative polymerase chain reaction. Kindlin-3 overexpression in a breast cancer cell line increased primary tumor growth and lung metastasis by 2.5- and 3-fold, respectively, when implanted into mice compared with cells expressing vector alone. Mechanistically, the Kindlin-3-overexpressing cells displayed a 2.2-fold increase in vascular endothelial growth factor (VEGF) secretion and enhanced ß1 integrin activation. Increased VEGF secretion resulted from enhanced production of Twist, a transcription factor that promotes tumor angiogenesis. Knockdown of Twist diminished VEGF production, and knockdown of ß1 integrins diminished Twist and VEGF production by Kindlin-3-overexpressing cells, while nontargeting small interfering RNA had no effect on expression of these gene products. Thus, Kindlin-3 influences breast cancer progression by influencing the crosstalk between ß1 integrins and Twist to increase VEGF production. This signaling cascade enhances breast cancer cell invasion and tumor angiogenesis and metastasis.

Tobacco smoke exposure and endothelial dysfunction in patients with advanced coronary artery disease.

INTRODUCTION:  Exposure to tobacco smoke is associated with a higher cardiovascular risk, especially in patients with coronary artery disease (CAD).  OBJECTIVES:  The aim of the study was to evaluate the effect of active and passive tobacco smoking on the activity of endothelial markers in advanced atherosclerosis. PATIENTS AND METHODS:  We studied 181 consecutive patients with advanced CAD (53 women and 128 men) aged 60 ±8 years, including 102 active self­declared smokers (56.3%). We determined plasma asymmetric dimethylarginine (ADMA), thrombomodulin (TM), and plasminogen activator inhibitor­1 (PAI­1) levels, along with serum cotinine concentrations as a marker of tobacco smoking.  RESULTS:  Plasma ADMA levels were higher in active smokers compared with nonsmokers (0.60 ±0.09 µmol/l vs. 0.49 ±0.08 µmol/l, P <0.001). There were similar intergroup differences in TM (4.60 ±2.11 ng/ml vs. 3.0 ±1.7 ng/ml, P <0.0001) and PAI­1 levels (30.3 ±12.4 ng/ml vs. 23.6 ±11.3 ng/ml, P <0.0001). We observed positive correlations between cotinine and ADMA (r = 0.71, P <0.0001), TM (r = 0.53, P <0.0001), and PAI­1 (r = 0.58, P <0.0001). In 21 patients (26.6%) who declared to be nonsmokers, cotinine levels (mean, 6.30 ±22.5 ng/ml) significantly correlated with ADMA, TM, and PAI­1 (all P <0.001). A multivariate regression analysis showed that cotinine was an independent predictor of ADMA, TM, and PAI­1 in the whole patient group.  CONCLUSIONS:  Despite long­lasting endothelial injury in advanced CAD, continued cigarette smoking is able to further enhance endothelial damage by increasing ADMA levels and resultant inhibition of fibrinolysis.

Kindlin-2 regulates hemostasis by controlling endothelial cell-surface expression of ADP/AMP catabolic enzymes via a clathrin-dependent mechanism.

Kindlin-2, a widely distributed cytoskeletal protein, has been implicated in integrin activation, and its absence is embryonically lethal in mice. In the present study, we tested whether hemostasis might be perturbed in kindlin-2(+/-) mice. Bleeding time and carotid artery occlusion time were significantly prolonged in kindlin-2(+/-) mice. Whereas plasma concentrations/activities of key coagulation/fibrinolytic proteins and platelet counts and aggregation were similar in wild-type and kindlin-2(+/-) mice, kindlin-2(+/-) endothelial cells (ECs) showed enhanced inhibition of platelet aggregation induced by adenosine 5'-diphosphate (ADP) or low concentrations of other agonists. Cell-surface expression of 2 enzymes involved in ADP/adenosine 5'-monophosphate (AMP) degradation, adenosine triphosphate (ATP) diphosphohydrolase (CD39) and ecto-5'-nucleotidase (CD73) were increased twofold to threefold on kindlin-2(+/-) ECs, leading to enhanced ATP/ADP catabolism and production of adenosine, an inhibitor of platelet aggregation. Trafficking of CD39 and CD73 at the EC surface was altered in kindlin-2(+/-) mice. Mechanistically, this was attributed to direct interaction of kindlin-2 with clathrin heavy chain, thereby controlling endocytosis and recycling of CD39 and CD73. The interaction of kindlin-2 with clathrin was independent of its integrin binding site but still dependent on a site within its F3 subdomain. Thus, kindlin-2 regulates trafficking of EC surface enzymes that control platelet responses and hemostasis.

[Osteopontin level in the serum of patients with alcoholic liver disease, treated for alcohol withdrawal syndrome--a preliminary report]. / Poziom osteopontyny w surowicy krwi u pacjentów z alkoholowa choroba watroby, leczonych z powodu alkoholowego zespolu odstawiennego--doniesienie wstepne.

Alcoholic liver disease (ALD) is consequence of alcohol addiction, that increase morbidity and mortality of those group of patients. Induction of the inflammatory process in alcohol injured liver is a turning point in ALD, can cause acute liver damage symptoms or cirrhosis with increase of hepatic fibrosis intensity and portal hypertension. Osteopontin (OPN)--a protein associated with early inflammatory process and remodeling of damaged tissues with intensification of hepatic fibrogenesis process, is considered to be a biomarker of hepatic inflammation and fibrosis. A group of 12 patients with mean age--44 years, mean alcohol addiction time--16 years, hospitalized because of alcohol withdrawal symptoms were examined in the preliminary report. ALD was diagnosed in all patients, steatosis--in 4, inflammatory--in 3, cirrhosis--in 5. Control group were 19 healthy patients with no alcohol addiction and liver disease in medical history. Serum OPN level was measured by ELISA method using reagents from Raybiotech. Serum OPN level in control group ranged 0-35,955 pg/ml, in examined group ranged 0-338,280 pg/ml. OPN levels in serum of patients with hepatic steatosis were under limit of detection (< 50 pg/ml). OPN serum level increase was correlated to ALD progression. The highest OPN levels (> 100,000 pg/ml) were observed in patients with symptoms of the liver decompensation due to severe hepatitis and advanced cirrhosis.

[Acute clozapine poisonings in years 2007 - 2010 in material of Clinic of Toxicology in Kraków]. / Ostre zatrucia klozapina w materiale Kliniki Toksykologii w Krakowie z lat 2007 - 2010.

18 patients with acute clozapine poisoning, 6 female and 12 male, were analyzed. The mean age was 42.8 years. Six patients were intoxicated only clozapine. Mixed poisoning (clozapine and other factor) was diagnosed in nine cases. Among the additional factors dominated psychotropic drugs. According to the Poisoning Severity Score (PSS) criteria in the study group was only a one mild intoxication. Acute pneumonia developed in 3 patients, acute bronchitis and rabdomyolysis were reported in one case. The most common symptoms included: agitation, confusion (83.3%), tachycardia (77.8%), CNS depression (66.7%), excessive mucus production in bronchi, hypersalivation (44.4%), miosis (50%). Disordered breathing requiring intubation or mechanical ventilation occurred in 27.7% of poisoned. The average duration of hospitalization was less than 7 days.

[Side effects of the HMG-CoA reductase inhibitors (statins). Lupus erythematosus induced by Atorvastatin therapy]. / Dzialania uboczne inhibitorów reduktazy HMG-co A (statyn). Indukowany przyjeciem atorvastatyny toczen rumieniowaty.

The paper describes the case of 56 years old woman admitted to the Toxicology Department because of skin lesions, joint and muscle pain and elevated activity of transaminases and creatine phosfokinase as well in biochemical analysis. The symptoms occurred after 6 days of the Atorvastatin therapy. The clinical picture indicated side effects of the hipolipemic therapy, but the presence of the skin lesions suggested drug induced collagenosis (lupus erythrematosus, dermatomyositis). Immunological studies confirmed association with antinuclear antibodies (ANA) and anti-Mi-2 autoantibodies in the serum. Immunosuppressive therapy was ordered with clinical and biochemical improvement.

[Sulpiride poisoning--case report confirmed with the quantitative determination of the xenobiotic serum level]. / Zatrucie sulpirydem--opis przypadku potwierdzony ilosciowym oznaczeniem stezenia ksenobiotyku w surowicy.

Despite above 40 years the presence of sulpride on the pharmaceutical market, the acute poisonings are poorly reported in the medical literature. The discussed case of sulpiride intoxication concerns ingestion probably dose of 12 g, that exceeded 10-fold maximum therapeutic dose. 16-year-old girl, with no previous sulpiride treatment, was admitted to the Toxicology Department about 3 hours after ingestion. In clinical picture she presented quantitative consciousness disturbances with maximum 10 scores in GCS scale, with tendency to low BP (minimum 88/45 mmHg) and episode of orthostatic hypotension. The ECG demonstrated: normogram, sinus tachycardia with a heart rate of 125 beats/min, PQ = 120 ms, QRS = 80 ms, prolongation of QTc to 519,6 ms and unspecific changes of ST-T syndrome. The qualitative toxicological test confirmed the presence of chlorprothixene in urine, but the serum therapeutic concentration (0.126 microg/ml) excluded the overdose. The quantitative determination of sulpiride serum concentration confirmed acute sulpiride poisoning. The measured sulpiride toxic concentration on admission and in the consecutive hours were from 13.2 to 8.2 microg/ml. Sulpiride toxicokinetic parameters such as t max = about 3 h, t 1/2 = 24.02 h, k(el) = 0.029 h(-1) were also estimated. They point out that the absorption rate is similar and the elimination is prorogated in sulpiride acute poisoning compared to therapeutic doses.

The integrin coactivator kindlin-2 plays a critical role in angiogenesis in mice and zebrafish.

Kindlin-2, a widely distributed cytoskeletal protein, has been implicated in integrin activation, and its absence is embryonically lethal in mice and causes severe developmental defects in zebrafish. Knockdown of kindlin-2 levels in endothelial cells resulted in defective adhesive and migratory responses, suggesting that angiogenesis might be aberrant even with partial reduction of kindlin-2. This hypothesis has now been tested in the kindlin-2(+/-) mice. RM1 prostate tumors grown in kindlin-2(+/-) mice had fewer blood vessels, which were thinner and shorter and supported less tumor growth compared with wild-type littermates. The vessels that did form in the kindlin-2(+/-) mice lacked smooth muscle cells and pericytes and had thinner basement membranes, indicative of immature vessels. VEGF-induced angiogenesis in matrigel implants was also abnormal in the kindlin-2(+/-) mice. Vessels in the kindlin-2(+/-) mice were leaky, and BM transplantation from kindlin-2(+/-) to WT mice did not correct this defect. Endothelial cells derived from kindlin-2(+/-) mice had integrin expression levels similar to WT mice but reduced αVß3-dependent signaling, migration, adhesion, spreading, and tube formation. Developmental angiogenesis was markedly impaired by kindlin-2 morpholinos in zebrafish. Taken together, kindlin-2 plays an important role in pathologic and developmental angiogenesis, which arises from defective activation of integrin αVß3.

Carbamazepine intoxication in alcohol dependent epileptic patients.

Carbamazepine is frequently administrated to alcohol addict patients. The aim of this study was to evaluate the influence of alcohol addiction on carbamazepine pharmacokinetics and severity of drug intoxication. The total of 158 carbamazepine intoxicated patients participated in the study (76 non-alcohol-dependent, and 82 alcohol-dependent subjects). The results of the study indicate that the level of unconsciousness depends on carbamazepine concentration. The frequency of anticholinergic toxidrome was higher in alcohol-dependent patients (88.6% - alcohol-abused subjects, and 78.3% - alcoholics in abstinency) comparing to non-alcohol-dependent epileptics (67.1%). The average biological half-life of carbamazepine in non-addicted epileptics was 41.5 h, 43.5 h for alcohol-dependent patients during abstinency, and 38.6 in abused patients. It may be concluded that ethanol doesn't influence the pharmacokinetic and pharmacodynamic of carbamazepine in acute drug intoxication.

[Amanitin determination in mushroom poisoning diagnostics]. / Oznaczanie amanityny w diagnostyce zatruc grzybami.

There are some serious poisonings with toxic mushroom species in Poland every year. Good prognostics in the cases is correlated to short time from mushroom consumption to hospitalization, correct distinguish not specific gastrointestinal and Amanita phalloides syndrome and immediately specific treatment. The purpose of the paper was to make appraisal of usefulness of amanitin blood and urine determination and transaminases activity determination (ALT, AST) in diagnostics of mushroom poisoned patients up to three days after mushroom consumption. The material was twenty two retrospective histories of mushroom poisoned patients treated in the years 2007-2008. Amanitin blood and urine determinations were made by ELISA method. Urine amanitin results in samples collected within 40 h from mushroom consuming were positive in all Amanita phalloides syndrome cases. Serum amanitin determination was not useful for the diagnostics. Trans-aminases activity determinations let to distinguish Amanita phalloides syndrome on the second and the third day after mushroom consumption. In the first poisoning phase (within 24 h), the ALT and AST activities were in normal ranges and only amanitin urine determination let to confirm or exclude Amanita phalloides poisoning. Amanitin urine determinations were useful to take fast decision about specific treatment and avoid internal organs dysfunctions.