Lenvatinib Plus Pembrolizumab in Previously Treated Advanced Endometrial Cancer: Updated Efficacy and Safety From the Randomized Phase III Study 309/KEYNOTE-775.

Clinical trials frequently include multiple end points that mature at different times. The initial report, typically based on the primary end point, may be published when key planned co-primary or secondary analyses are not yet available. Clinical Trial Updates provide an opportunity to disseminate additional results from studies, published in JCO or elsewhere, for which the primary end point has already been reported.We report the final prespecified analysis for overall survival (OS), along with updated progression-free survival (PFS) and objective response rate (ORR), and safety from the open-label, randomized, phase III Study 309/KEYNOTE-775. In total, 827 patients with advanced, recurrent, or metastatic endometrial cancer (EC) were randomly assigned to receive lenvatinib 20 mg orally once daily plus pembrolizumab 200 mg intravenously once every 3 weeks (n = 411) or chemotherapy of the treating physician's choice (doxorubicin 60 mg/m2 intravenously once every 3 weeks or paclitaxel 80 mg/m2 intravenously once weekly [3 weeks on; 1 week off] [n = 416]). Efficacy was reported for patients with mismatch repair proficient (pMMR) tumors and all-comers, and by subgroups (histology, prior therapy, MMR status). Updated safety was also reported.Lenvatinib plus pembrolizumab showed benefits in OS (pMMR HR, 0.70; 95% CI, 0.58 to 0.83; all-comer HR, 0.65; 95% CI, 0.55 to 0.77), PFS (pMMR HR, 0.60; 95% CI, 0.50 to 0.72; all-comer HR, 0.56; 95% CI, 0.48 to 0.66), and ORR (pMMR patients, 32.4% v 15.1%; all-comers, 33.8% v 14.7%) versus chemotherapy. OS, PFS, and ORR favored lenvatinib plus pembrolizumab in all subgroups of interest. No new safety signals were observed. Lenvatinib plus pembrolizumab continued to show improved efficacy versus chemotherapy and manageable safety in patients with previously treated advanced EC.

Lenvatinib plus Pembrolizumab for Advanced Endometrial Cancer.

BACKGROUND: Standard therapy for advanced endometrial cancer after failure of platinum-based chemotherapy remains unclear. METHODS: In this phase 3 trial, we randomly assigned, in a 1:1 ratio, patients with advanced endometrial cancer who had previously received at least one platinum-based chemotherapy regimen to receive either lenvatinib (20 mg, administered orally once daily) plus pembrolizumab (200 mg, administered intravenously every 3 weeks) or chemotherapy of the treating physician's choice (doxorubicin at 60 mg per square meter of body-surface area, administered intravenously every 3 weeks, or paclitaxel at 80 mg per square meter, administered intravenously weekly [with a cycle of 3 weeks on and 1 week off]). The two primary end points were progression-free survival as assessed on blinded independent central review according to the Response Evaluation Criteria in Solid Tumors, version 1.1, and overall survival. The end points were evaluated in patients with mismatch repair-proficient (pMMR) disease and in all patients. Safety was also assessed. RESULTS: A total of 827 patients (697 with pMMR disease and 130 with mismatch repair-deficient disease) were randomly assigned to receive lenvatinib plus pembrolizumab (411 patients) or chemotherapy (416 patients). The median progression-free survival was longer with lenvatinib plus pembrolizumab than with chemotherapy (pMMR population: 6.6 vs. 3.8 months; hazard ratio for progression or death, 0.60; 95% confidence interval [CI], 0.50 to 0.72; P<0.001; overall: 7.2 vs. 3.8 months; hazard ratio, 0.56; 95% CI, 0.47 to 0.66; P<0.001). The median overall survival was longer with lenvatinib plus pembrolizumab than with chemotherapy (pMMR population: 17.4 vs. 12.0 months; hazard ratio for death, 0.68; 95% CI, 0.56 to 0.84; P<0.001; overall: 18.3 vs. 11.4 months; hazard ratio, 0.62; 95% CI, 0.51 to 0.75; P<0.001). Adverse events of grade 3 or higher occurred in 88.9% of the patients who received lenvatinib plus pembrolizumab and in 72.7% of those who received chemotherapy. CONCLUSIONS: Lenvatinib plus pembrolizumab led to significantly longer progression-free survival and overall survival than chemotherapy among patients with advanced endometrial cancer. (Funded by Eisai and Merck Sharp and Dohme [a subsidiary of Merck]; Study 309-KEYNOTE-775 ClinicalTrials.gov number, NCT03517449.).

Development and inter-laboratory validation of the VISAGE enhanced tool for age estimation from semen using quantitative DNA methylation analysis.

The analysis of DNA methylation has become an established method for chronological age estimation. This has triggered interest in the forensic community to develop new methods for age estimation from biological crime scene material. Various assays are available for age estimation from somatic tissues, the majority from blood. Age prediction from semen requires different DNA methylation markers and the only assays currently developed for forensic analysis are based on SNaPshot or pyrosequencing. Here, we describe a new assay using massively parallel sequencing to analyse 13 candidate CpG sites targeted in two multiplex PCRs. The assay has been validated by five consortium laboratories of the VISible Attributes through GEnomics (VISAGE) project within a collaborative exercise and was tested for reproducible quantification of DNA methylation levels and sensitivity with DNA methylation controls. Furthermore, DNA extracts and stains on Whatman FTA cards from two semen samples were used to evaluate concordance and mimic casework samples. Overall, the assay yielded high read depths (> 1000 reads) at all 13 marker positions. The methylation values obtained indicated robust quantification with an average standard deviation of 2.8% at the expected methylation level of 50% across the 13 markers and a good performance with 50 ng DNA input into bisulfite conversion. The absolute difference of quantifications from one participating laboratory to the mean quantifications of concordance and semen stains of remaining laboratories was approximately 1%. These results demonstrated the assay to be robust and suitable for age estimation from semen in forensic investigations. In addition to the 13-marker assay, a more streamlined protocol combining only five age markers in one multiplex PCR was developed. Preliminary results showed no substantial differences in DNA methylation quantification between the two assays, indicating its applicability with the VISAGE age model for semen developed with data from the complete 13-marker tool.

Intramyocardial Transplantation of Umbilical Cord Mesenchymal Stromal Cells in Chronic Ischemic Cardiomyopathy: A Controlled, Randomized Clinical Trial (HUC-HEART Trial).

BACKGROUND AND OBJECTIVES: The HUC-HEART Trial (ClinicalTrials.gov Identifier: NCT02323477) was a controlled, prospective, phase I/II, multicenter, single-blind, three-arm randomized study of intramyocardial delivery of human umbilical cord-derived mesenchymal stromal cells (HUC-MSCs) combined with coronary artery bypass-grafting (CABG) in patients with chronic ischemic cardiomyopathy (CIC). The trial aimed to assess (i) the safety and the efficacy of cell transplantation during one-year follow-up, (ii) to compare the efficacy of HUC-MSCs with autologous bone-marrow- derived mononuclear cells (BM-MNCs) in the same clinical settings. METHODS AND RESULTS: Fifty-four patients who were randomized to receive HUC-MSCs (23×106) (n=26) or BM-MNCs (70×107) (n=12) in combination with CABG surgery. The control patients (n=16) received no cells/vehicles but CABG intervention. All patients were screened at baseline and 1, 3, 6, 12 months after transplantation. Forty-six (85%) patients completed 12 months follow-up. No short/mid-term adverse events were encountered. Decline in NT-proBNP (baseline∼ 6 months) in both cell-treated groups; an increase in left ventricular ejection fraction (LVEF) (5.4%) and stroke volume (19.7%) were noted (baseline∼6 or 12 months) only in the HUC-MSC group. Decreases were also detected in necrotic myocardium as 2.3% in the control, 4.5% in BM-MNC, and 7.7% in the HUC-MSC groups. The 6-min walking test revealed an increase in the control (14.4%) and HUC-MSC (23.1%) groups. CONCLUSIONS: Significant findings directly related to the intramyocardial delivery of HUC-MSCs justified their efficacy in CIC. Stricter patient selection criteria with precisely aligned cell dose and delivery intervals, rigorous follow-up by detailed diagnostic approaches would further help to clarify the responsiveness to the therapy.

Umbilical cord mesenchymal stromal cells engraft and transdifferentiate into cardiomyocyte-like cells following acute myocardial ischemia⋆.

OBJECTIVE: Human umbilical cord-derived mesenchymal stromal cells (hUC-MSCs) gained importance in acute/chronic ischemic cardiomyopathy because of their outstanding regenerative potential in various pathologic conditions. The present study was designed to determine to what extent hUC-MSCs contribute to myocardial regeneration in acute experimental myocardial infarction (MI) in rats. METHODS: Animals were assigned into two groups; the control group received intramyocardial PBS injections, while the hUC-MSC group received calcein-AM-labeled 8.8 × 106/kg hUC-MSCs. Three weeks following the acute MI induction, rats were sacrificed after assessing the left ventricular (LV) function using echocardiography. For the assessment of infarct size, the triphenyl tetrazolium chloride (TTC) test was used in isolated hearts. Collagen-rich scar tissue was demonstrated using Masson's trichrome staining, followed by the detection of cardiac troponin I (cTnI), α-sarcomeric actin (α-SA), von Willebrand factor (vWF), CD68 and CD206 expressions in control and cell-injected sections. RESULTS: Echocardiography revealed a significant difference (P = 0.037) in the LV ejection fraction between groups. TTC assays demonstrated a significant difference (P = 0.006) between the groups regarding the ratio of the infarcted LV area. Calcein-AM-loaded cells were identified mostly in ischemic myocardium. Transplanted cells also expressed human-specific cTnI, providing concrete proof of transdifferentiation into cardiomyocytes, and α-SA. vWF+ cells verified the neovascularization in the ischemic myocardium. Finally, a slight shift from pro-inflammatory to anti-inflammatory macrophages (CD68+/CD206+) was noted in both groups. CONCLUSIONS: We found that the intramyocardial transplanted hUC-MSCs engrafted and partially transdifferentiated into cardiomyocytes, reduced scar formation, and induced angiogenesis through the association of pro/anti-inflammatory macrophages.

Body fluid identification and assignment to donors using a targeted mRNA massively parallel sequencing approach - results of a second EUROFORGEN / EDNAP collaborative exercise.

In a previous EUROFORGEN/EDNAP collaborative exercise, we tested two assays for targeted mRNA massively parallel sequencing for the identification of body fluids/tissues, optimized for the Illumina MiSeq/FGx and the Ion Torrent PGM/S5 platforms, respectively. The task of the second EUROFORGEN/EDNAP collaborative exercise was to analyze dried body fluid stains with two different multiplexes, the former Illumina 33plex mRNA panel for body fluid/tissue identification and a 35plex cSNP panel for assignment of body fluids/tissues to donors that was introduced in a proof-of-concept study recently. The coding region SNPs (cSNPs) are located within the body fluid specific mRNA transcripts and represent a direct link between the body fluid and the donor. We predicted the origin of the stains using a partial least squares discriminant analysis (PLS-DA) model, where most of the single source samples were correctly predicted. The mixed body fluid stains showed poorer results, however, at least one component was predicted correctly in most stains. The cSNP data demonstrated that coding region SNPs can give valuable information on linking body fluids/tissues with donors in mixed body fluid stains. However, due to the unfavorable performance of some cSNPs, the interpretation remains challenging. As a consequence, additional markers are needed to increase the discrimination power in each body fluid/tissue category.

Optimizing the transport and storage conditions of current Good Manufacturing Practice -grade human umbilical cord mesenchymal stromal cells for transplantation (HUC-HEART Trial).

BACKGROUND: The HUC-HEART Trial is a clinical study of intramyocardial delivery of current Good Manufacturing Practice (cGMP)-grade human umbilical cord multipotent stromal cells (HUC-MSCs) in ischemic cardiomyopathy where 2 × 107 cells are administered to peri-infarcted myocardium. Prior to the onset of the trial, we aimed to optimize the transport/storage conditions for obtaining the highest cell viability and proliferation rate of cells to be transplanted. METHODS: Cells were tested after being transported in phosphate-buffered saline (PBS) or Ringer's lactate-based (RL) transport media supplemented with human serum albumin (HSA) and/or hydroxyethyl starch (HES) at two temperatures (2-10°C or 22-24°C). RESULTS: The effects of transport conditions on cell viability following 6 h were found highest (93.4 ± 1.5) in RL-based media at 2-10°C. Karyotypes were found normal upon transportation in any of the formulations and temperatures. However, the highest proliferation rate was noted (3.1-fold increase) in RL (1% HSA) media at 2-10°C over 6 days in culture. From that point, RL (1% HSA) media at 2-10°C was used for further experiments. The maximum cell storage time was detected around 24 h at 2-10°C. Extended storage periods resulted in a decrease in cell viability but not in MSC marker expression. An increase in actin quantity was detected in hypoxia (5% O2) groups in early culture days; no difference was noted between hypoxic versus normoxic (21% O2) conditions in later days. DISCUSSION: The overall results suggest that non-commercial, simple media formulations with extended storage intervals at 2-10°C temperatures are capable of retaining the characteristics of clinical-grade HUC-MSCs. The above findings led us to use RL (1% HSA) media at 2-10°C for transport and storage in the HUC-HEART Trial; 23 patients received HUC-MSCs by August 2018; no adverse effects were noted related to cell processing and transplantation.

Comparison of the Ovary and Kidney as Sites for Islet Transplantation in Diabetic Rats.

BACKGROUND: Currently, the most commonly used site for clinical islet transplantation is the liver although it is far from being an ideal site. Low oxygen tension and the induction of an inflammatory response impair islet implantation and lead to significant early loss of islet. The present study aimed to investigate and compare the efficacy of islet transplantation to the ovary and kidney subcapsule in diabetic rats. METHODS: The study was performed with 3 groups of rats (control, ovary, and kidney subcapsule) including 6 Sprague female rats each. Diabetes model was created with the use of streptozotocin, and blood glucose levels of the rats were measured after 72 hours. Thirty days after the transplantation, blood samples were obtained from the rats, and then pancreas, kidney, and ovary specimens were fixed in 10% formaldehyde and the experiment completed. After staining with hematoxylin and eosin, the tissue samples were morphologically evaluated by a specialist histopathologist. RESULTS: Changes in mean blood glucose and C-peptide levels were statistically significant in the ovary and kidney subcapsule groups. Histologic examination revealed that granulosus insulin-bearing cells were detected in the islet grafts of both ovary and kidney subcapsule groups. The renal subcapsule group had inflammation signs on histologic examination. The islet cells of both ovary and renal subcapsule groups had no vacuolization. CONCLUSIONS: We showed that the ovary might be a new site for islet transplantation. Further research should be done on whether the initial results of this study can be reproduced in larger numbers of animal models and eventually in humans.

Which type of conditioning method protects the spinal cord from the ischemia-reperfusion injury in 24 hours?

OBJECTIVE: This study was designed to test the effects of different types of preconditioning and postconditioning methods on spinal cord protection following aortic clamping. METHODS: The animals (rabbits) were divided into sham-operated, ischemic preconditioning, remote ischemic preconditioning, simultaneous aortic and ischemic remote preconditioning, and ischemic postconditioning groups. After neurological evaluations, ultrastructural analysis and immunohistochemical staining for caspase-3 were evaluated after 24 h following ischemia. RESULTS: The neurological outcomes of the remote ischemic preconditioning (4.2 ± 0.4) and ischemic postconditioning (4.6 ± 0.8) groups were significantly improved when compared with the ischemia group (2.2 ± 04). The immunohistochemical analysis revealed that the lowest percentage of apoptosis was in-group ischemic preconditioning at 12.5 ± 30.6%. In the comparison of intracellular edema in an ultrastructural analysis, the ischemic preconditioning and ischemic postconditioning groups had significantly lower values than the ischemia group. CONCLUSION: The conditioning methods attenuate ischemia-reperfusion injury for spinal cord injury. Ischemic and remote preconditioning and also postconditioning methods are simple to perform and inexpensive.

The effect of CYP1A1 and GSTP1 isozymes on the occurrence of aortic aneurysms.

BACKGROUND: Aortic aneurysms are vascular diseases that are associated with high mortality and morbidity. Cytochrome P450 CYP1A1 and glutathione S-transferase (GSTP1) isozymes were searched and compared with the patients who had experienced aortic surgery due to aortic aneurysm and atherosclerotic patients without aneurysm to find the relation of the oxidative stress with the aneurysms. MATERIALS AND METHODS: Study group consisted of the patients with the diagnosis of aortic aneurysm (group I, n: 12) and control group who were operated for coronary bypass surgery: preoperatively drug users (group II, n: 21) and nonusers (group III, n: 15). Paraffin sections (4 µm thick) of aortic biopsy materials were stained with hematoxylin and eosine, CYP1A1 and GSTP1 immunohistochemical markers. The specimens were evaluated using light microscopy at 40- to 400-fold magnification. RESULTS: The expressions of CYP1A1 and GSTP1 isozymes were found statistically significantly higher in the patients who have an aortic aneurysm than both the control groups (p < 0.05). There was no significant association between protein expressions, drugs and duration of usage, patient's demographic variables, and smoking (p > 0.05). CONCLUSIONS: In this pioneering study, CYP1A1 and GSTP1 isozymes are related with the aneurysms. The strategy that prevents the oxidative stress for the patients who had aortic aneurysms could be a valuable choice of searching to effect the aneurysmal progression.

Ruptured abdominal aortic aneurysm treatment in the stent graft era.

BACKGROUND: We aim to decrease mortality and morbidity by early diagnosis and endovascular aneurysm repair (EVAR) or by using open surgery. METHODS: The patients who had underwent open surgery and EVAR with a diagnosis of ruptured abdominal aortic aneurysms were evaluated retrospectively. Patients with EVAR were separated as group I and the patients with surgical operations constituted group II. The risk factors, duration of the operation, blood product usage, drainage amounts, complications, mortality, and morbidity rates were evaluated. RESULTS: The duration of the operation and the required blood and blood products were lower in group I (P < .05). There is no any significant difference between the groups in terms of mortality, complications, short-, and long-term results. CONCLUSION: We support the idea that better results can be obtained by showing regard to suitable patient, suitable clinical condition, and suitable anatomy together with the correct choice of operation type.

Effect of conditioning on visceral organs during indirect ischemia/reperfusion injury.

BACKGROUND: The mortality and morbidity rates of even extensive thoracoabdominal replacement have improved markedly in recent years. We investigated the effects of a temporary occlusion of the aorta as a direct precondition and temporary occlusion of the axillary artery for remote preconditioning to determine any effects that preconditioning may have on indirect (nonischemic) injuries to visceral organs (indirect effects of remote ischemia/reperfusion injury). METHODS: Thirty-seven New Zealand white rabbits were divided into five groups: controls (sham-operated; group 1); direct ischemia to the infrarenal aorta without preconditioning (group 2); direct ischemic preconditioning to the infrarenal aorta (group 3); remote ischemic preconditioning before clamping the infrarenal aorta (group 4); and simultaneous direct aortic and remote ischemic preconditioning before the clamping and during clamping of the infrarenal aorta (group 5). We used a 30-minute ischemia period for aortic occlusion for spinal cord ischemia/reperfusion. The axillary artery was used for remote preconditioning. After 24 hours, tissue specimens of the internal organs were obtained. RESULTS: Myocardial congestion was the main pathology detected in all groups. Histopathologic evaluation of tissue samples taken from the hearts showed no significant differences in terms of the degree of polymorphonuclear leukocyte (PMNL) infiltration and edema between the groups. Lung congestion and pneumonic cell infiltration were detected in all the groups. Pneumonic cell infiltration was significantly high in groups 2 and 3. Cell infiltration was lowest in group 4 at 71.4% of normal values, which differed from the normal values of 25-33.3% in the other groups (P < 0.05). Although there is a difference between the groups in case of renal congestion, there is not any difference as tubular damage and PMN. There was a significant difference with regard to renal congestion between groups 2 and 3. Renal congestion was normal in 80% of the kidneys in group 3. This differed from the normal values observed in the other groups (14.3-57.1%, P < 0.05). Liver congestion was detected in all groups. CONCLUSIONS: Different preconditioning methods may play an important role in distinct organ injuries during aortic cross-clamping. The visceral organs that exhibited positive and constructive results with direct and remote preconditioning included the lungs and kidneys during indirect ischemia/reperfusion injury. Remote ischemic conditioning was determined to be especially advantageous as a protection method, due to the fact that it is easy to use and effective for indirect ischemia/reperfusion injury.

The contralateral extremity has also benefit from the locally administered bone marrow-derived mononuclear cells and cord blood serum in diabetic ischemic wound healing.

OBJECTIVE: Impaired wound healing could be a disaster especially in diabetes and amputation is the major risk. The aim of this study was to evaluate the benefit of BMMCs and CBS on wound healing. METHODS: Diabetic rats were underwent bilateral limb ischemia and wounding of skin defects on both extremities. The groups were formed as BMMCs (group A), BMMCs and CBS (group B), only CBS (group C), and phosphate buffer solution (group D) that were injected into wounds on right legs. RESULTS: The complete recovery of right legs was established as a mean of 21.4 ± 1.1 days, 12.9 ± 1.5 days, 30.0 ± 0.0 days and 38.1 ± 1.5 days according to Groups A, B, C, and D (p < 0.05). The recovery of left legs were calculated as a mean of 27.0 ± 0.0 days, 24.0 ± 0.0 days, 35.6 ± 1.1 days and 37.3 ± 1.6 days according to Groups A, B, C and D (p < 0.05). At the end of the recovery, the HE staining showed that vascularity was increased in groups A and B. CONCLUSION: Transplantation of BMMCs and CBS to the ischemic wounds of the diabetic rats accelerate the repair. The recovery was also superior in the same group although the treatment was not applied to the left extremity directly.

Combined genetic mutations have remarkable effect on deep venous thrombosis and/or pulmonary embolism occurence.

PURPOSE: Although deep vein thrombosis and thromboembolic diseases differ among various races, they are still important in our day. The difficulties in treatment and following-up of these diseases are caused by secret genetic mutations rather than predisposing factors. METHODS: Between January 2011 and May 2013, patients who were traced for deep vein thrombosis and/or pulmonary embolism were evaluated retrospectively. 84 patients (53.6% males and 46.4% females) were included in the study. Their family histories, predisposing factors and treatments were researched. Factor V Leiden (G 1691A), Factor II G20210A, Plasminogen Activator Inhibitor-Type 1 (4G/5G), and Methylene Tetrahydrofolate Reductase (C677T, A1298C) mutations were investigated from peripheral venous blood. RESULTS: Among the genetic mutations we searched, the incidence of single mutation rate was observed at 11.9%, double mutation collocation at 44%, triple mutation collocation at 29.8%, quadruple mutation collocation at 13.1%, and finally, quintuplet mutation collocation at 1.2%. Our approximate mutation number was found as 2.47 ± 0.91. CONCLUSION: We observed that multiple mutations were high in number compared to single genetic mutations. The patients who have multiple mutations should be more in the front line considering their diagnosis, treatment and following up, and also in terms of decreasing mortality, morbidity and recurrence.

Right ventricular and septal function in patients with pulmonary hypertension.

OBJECTIVE: Pulmonary hypertension (PHT) exacerbates the functions of both ventricles. This prospective, randomised study was planned to investigate the effects of PHT on kinetics of both ventricles and the septum. METHODS: Twenty-five patients were randomly selected among the patients who had been planned to undergo mitral valve replacement (MVR) because of isolated mitral stenosis and divided into two groups according to their preoperative pulmonary artery pressure (PAP) values. Blood pool gated single photon emission tomography (BPGS) and transthoracic echocardiography were performed. Ventricles' regional, global and functional parameters were also assessed by using pulsed wave Doppler tissue imaging (DTI). RESULTS: Preoperative and postoperative PAP of the group 1 (PAP < 50 mmHg) were 40.0 ± 2.8 and 30.0 ± 2.6 mmHg (p = 0.03), group 2 (PAP ≥ 50 mmHg) were 71.9 ± 4.7 and 50.6 ± 3.5 mmHg (p < 0.05). The global right and left ventricle scores were decreased after the operation. The decrement was only significant in group 2. Considering the septal kinetics, right ventricle septal score was decreased from 7.6 to 3.3 (p < 0.05) in group 1, from 3.8 to 1.6 (p < 0.05) in group 2 postoperatively. CONCLUSION: Following MVR, a decrement in PAP values, and an improvement in ventricular function, especially in the right ventricular and septal kinetics were achieved. Furthermore, it was found that both DTI and BPGS techniques are beneficial to investigate the functional changes postoperatively and in the follow-up period of the patients who undergo mitral valve surgery.

Treatment efficacy with bone marrow derived mesenchymal stem cells and minocycline in rats after cerebral ischemic injury.

OBJECTIVE: We aimed to investigate the effects of bone marrow derived mesenchymal stem cells (MSCs), minocycline, and these two therapies combined on functional and histological improvement in cerebral ischemic injury created rats. MATERIALS AND METHODS: Twenty-eight Sprague Dawley female rats, weighing 250-300 g, were included in the study. Two male rats with similar properties were sacrificed for bone marrow derived MSC production. Group 1 was established as the control group. Group 2 was the group of only minocycline administered rats. Group 3 was the one of only MSCs administered rats. Group 4 was composed of the rats given the combination of MSCs and minocycline. Hematoxylin and eosin staining was done to assess the degeneration of the cells. Immunohistochemical staining was performed to evaluate the regeneration. Motor functions were examined by using Bederson's score. RESULTS: Cell degeneration was the least in group 4. The cells stained with GFAP were observed mostly in group 4. The cells stained with Neu N in group 1 were statistically lower than in other groups. When the groups were ordered in terms of functional improvement at the end of the second week, group 4 had the most and group 1 had the least. CONCLUSIONS: Bone marrow derived MSCs can lead to more histological and functional improvement when administered with minocycline, which is a neuroprotective agent as early as 24 h following the ischemic injury in a rat model. Minocycline therapy alone can be as effective as bone marrow derived MSCs therapy alone in ischemic cerebral rat model.

Influence of blood collection systems on coagulation tests.

OBJECTIVE: Coagulation tests are influenced by pre-analytic conditions such as blood collection systems. Change of glass collection tubes with plastic ones will cause alteration of the test results. The aim of this study was to compare three plastic blood collection tubes with a standard glass blood collection tube and each plastic collection tube with the other two for possible additional tube-to- tube differences. MATERIAL AND METHODS: A total of 284 blood samples were obtained from 42 patients receiving warfarin during their routine controls, besides 29 healthy volunteers. Subgroup analyses were done according to health status. RESULTS: Our study demonstrated that different blood collection tubes have a statistically significant influence on coagulation tests. The magnitude of the effect depends on the tube used. However most of the tests performed on samples obtained from any tube correlated significantly with results obtained from other tube samples. CONCLUSION: Although blood collection tubes with different brands or properties will have distinct effects on coagulation tests, the influence of these blood collection tubes may be relatively small to interfere with decision-making on dose prescription, therefore lack clinical importance. Correlations between the results showed that, one of these plastic blood collection tubes tested in our study, can be used interchangably for a wide variety of coagulation assays. CONFLICT OF INTEREST: None declared.

Relation between the frequency of CD34⁺ bone marrow derived circulating progenitor cells and the number of diseased coronary arteries in patients with myocardial ischemia and diabetes.

BACKGROUND: Bone marrow-derived circulating progenitor cells (BM-CPCs) in patients with coronary heart disease are impaired with respect to number and mobilization. However, it is unknown whether the mobilization of BM-CPCs depends on the number of diseased coronary arteries. Therefore, in our study, we analysed the correlation between the diseased coronary arteries and the frequency of CD34/45+ BM-CPCs in peripheral blood (PB) in patients with ischemic heart disease (IHD). METHODS: The frequency of CD34/45+ BM-CPCs was measured by flow cytometry in 120 patients with coronary 1 vessel (IHD1, n = 40), coronary 2 vessel (IHD2, n = 40), coronary 3 vessel disease (IHD3, n = 40) and in a control group of healthy subjects (n = 40). There was no significant difference of the total number of cardiovascular risk factors between IHD groups, beside diabetes mellitus (DM), which was significantly higher in IHD3 group compared to IHD2 and IHD1 groups. RESULTS: The frequency of CD34/45+ BM-CPCs was significantly reduced in patients with IHD compared to the control group (CD34/45+; p < 0.001). The frequency of BM-CPCs was impaired in patients with IHD3 compared to IHD1 (CD34/45+; p < 0.001) and to IHD2 (CD34/45+; p = 0.001). But there was no significant difference in frequency of BM-CPCs between the patients with IHD2 and IHD1 (CD34/45+; p = 0.28). In a subgroup we observed a significant negative correlation between levels of hemoglobin AIc (HbAIc) and the frequency of BM-CPCs (CD34/45+; p < 0.001, r = -0.8). CONCLUSIONS: The frequency of CD34/45+ BM-CPCs in PB is impaired in patients with IHD. This impairment may augment with an increased number of diseased coronary arteries. Moreover, the frequency of CD34/45+ BM-CPCs in ischemic tissue is further impaired by diabetes in patients with IHD.