Beyond brain injury biomarkers: chemoattractants and circulating progenitor cells as biomarkers of endogenous rehabilitation effort in preterm neonates with encephalopathy.

Introduction: Preclinical work and studies in adults have shown that endogenous regeneration efforts that involve mobilization of progenitor cells take place after brain injury. However, kinetics of endogenous circulating progenitor cells (CPCs) in preterm neonates is not well described, particularly their possible role regarding brain injury and regeneration. We aimed to assess the kinetics of CPCs in neonates with encephalopathy of prematurity in relation to brain injury biomarkers, chemoattractants and relevant antenatal and postanal clinical factors, in an effort to outline the related pathophysiology. Materials and methods: 47 preterm neonates (of 28-33 weeks GA) were enrolled: 31 newborns with no or minimal brain injury (grade I IVH) and 16 prematures with encephalopathy (grade III or IV IVH, PVL or infarct). Peripheral blood samples obtained on days 1, 3, 9, 18 and 45 after birth were analyzed using flow cytometry, focusing on EPCs (early and late Endothelial Progenitor Cells), HSCs (Hematopoietic Stem Cells) and VSELs (Very Small Embryonic-Like Stem Cells). At the same time-points serum levels of S100B, Neuron-specific Enolase (NSE), Erythropoietin (EPO), Insulin-like growth factor-1 (IGF-1) and SDF-1 were also measured. Neonates were assessed postnatally with brain MRI, and with Bayley III developmental test at 2 years of corrected age. Results: Preterms with brain injury proved to have significant increase of S100B and NSE, followed by increase of EPO and enhanced mobilization mainly of HSCs, eEPCs and lEPCs. IGF-1 was rather decreased in this group of neonates. IGF-1 and most CPCs were intense decreased in cases of antenatal or postnatal inflammation. S100B and NSE correlated with neuroimaging and language scale in Bayley III test, providing good prognostic ability. Conclusion: The observed pattern of CPCs' mobilization and its association with neurotrophic factors following preterm brain injury indicate the existence of an endogenous brain regeneration process. Kinetics of different biomarkers and associations with clinical factors contribute to the understanding of the related pathophysiology and might help to early discriminate neonates with adverse outcome. Timely appropriate enhancement of the endogenous regeneration effort, when it is suppressed and insufficient, using neurotrophic factors and exogenous progenitor cells might be a powerful therapeutic strategy in the future to restore brain damage and improve the neurodevelopmental outcome in premature infants with brain injury.

Iodine-lithium-alpha-dextrin (ILαD) against Staphylococcus aureus skin infections: a comparative study of in-vitro bactericidal activity and cytotoxicity between ILαD and povidone-iodine.

BACKGROUND: As antimicrobial resistance continues to increase, revisiting old antimicrobial agents, modified to enhance efficacy and safety, becomes important. Iodine has been widely used for more than 150 years as a wound and skin disinfectant; it is an effective broad range bactericide and does not promote the development of resistant strains. The most important iodine-based agent is povidone-iodine (PVP-I) which provides excellent antibacterial activity. However, its safety profile has been questioned. AIM: To evaluate the in-vitro antibacterial efficacy and kinetic properties of a novel iodine-based compound, iodine lithium alpha-dextrin (ILαD), against Staphylococcus aureus, and compare the in-vitro cytotoxicity profiles of ILαD and PVP-I. METHODS: A minimum inhibitory concentration (MIC) microbroth dilution method was performed against 12 meticillin-resistant (MRSA) and eight meticillin-susceptible (MSSA) S. aureus clinical isolates using ILαD and PVP-I. Time-kill and post-antibiotic effect studies of ILαD provided rate-of-kill information. MTT cytotoxicity assays were performed using three cell lines, treated with MIC doses of ILαD and PVP-I. FINDINGS: The MIC values of ILαD and PVP-I against the MRSA strains were 125 mg/L and 31.25 mg/L, respectively. Time-kill and post-antibiotic effect studies of ILαD revealed a log10 reduction factor of 3 within 8 h of exposure at a 2 × MIC dose; the post-antibiotic effect was calculated at 5±0.3h. Cell viability was affected slightly at the MIC dose of ILαD, while the MIC dose of PVP-I exerted a strong cell growth inhibitory effect of 90-95%. CONCLUSIONS: ILαD could be a promising solution against staphylococcal infections as it is effective, does not promote the development of resistant strains, and in-vitro testing indicates that it may be safer than PVP-I. Further studies are justified to determine whether ILαD overcomes the clinical limitations of PVP-I.

Matrix metalloproteinase-1 and -2 as markers of mineral bone disease in chronic kidney disease patients.

BACKGROUND: In the past few years, a distinct and multifactorial clinical entity called chronic kidney disease-mineral and bone disorder (CKD-MBD) that leads to decreased bone density and osteoporosis has been identified. The aim of this study was to assess the levels of the matrix metalloproteinase-1 and -2 (MMP-1 and MMP-2) in chronic kidney disease (CKD) patients of various disease stages in correlation to other bone turnover markers (BTM). This study is an initial investigative approach to a possible role of matrix metalloproteinases (MMPs) in the evaluation of bone disease in uremic patients. METHODS: We enrolled 60 patients at different stages of pre-dialysis CKD, 20 patients on hemodialysis (HD), and 20 age-matched healthy controls. Serum intact parathyroid hormone (iPTH), osteocalcin (OC), N-terminal propeptide of type I collagen (P1NP), and beta-C-terminal telopeptide of type I collagen (ß-CTX), were measured by electrochemiluminescence on automatic analyzers. Serum MMP-1 and MMP-2 levels were estimated using a commercial enzyme-linked immunosorbent assay (ELISA). Serum levels of urea, creatinine, calcium, phosphorus, and alkaline phosphatase were estimated. Creatinine clearance (ClCr) was calculated using the traditional clearance formula based on a 24-hour urine collection. RESULTS: Serum iPTH, OC, P1NP, ß-CTX concentrations were significantly higher (p <0.0001) while ClCr was significantly lower (p <0.0001) in CKD patients, as compared with those of healthy controls. A positive correlation was established between serum MMP-1 and OC levels (r =0.245, p =0.014), as well as with serum ß-CTX levels (r =0.197, p =0.048), and a negative correlation between MMP-2 and OC (r =-0.222, p =0.025). CONCLUSIONS: In CKD patients MMP-1 serum levels may reflect increased bone turnover rates. HIPPOKRATIA 2017, 21(1): 25-31.

Successful management of an equine carpal chip fracture by intra-articularly injected adipose-derived stromal vascular fraction after arthroscopic removal.

Carpal chip fractures are common causes of lameness in racehorses. Due to disadvantages in surgical management, adjuvant treatment modalities are usually necessary. Adipose-derived stem cells (ADSCs) have the potential to differentiate into other cell types including bone and cartilage cells. Adipose-derived stromal vascular fraction (SVF) is produced during ADSCs isolation from adipose tissue. The purpose of this report was to present the successful management of a grade III chip fracture in the right carpus of a 5-year-old Thoroughbred gelding by intra-articularly injected autologous SVF one month after the arthroscopic removal of the fracture. This treatment resulted in lameness improvement and short rehabilitation period to previous racing activities. High performance levels and no recurrent injuries were recorded during a twenty month follow-up period.

Mobilization of circulating progenitor cells following brain injury in premature neonates could be indicative of an endogenous repair process. A pilot study.

BACKGROUND: Preclinical data and adult studies have showed an endogenous regeneration process following brain damage that involves mobilization of progenitor cells. This process is not well described in preterm neonates. The present study aims to investigate the mobilization of Circulating Progenitor Cells (CPCs) and their relation to biomarkers of brain injury in preterm neonates. METHODS: This is a prospective cohort study of preterm infants with gestational age (GA) <34 weeks. Serial cranial ultrasounds scans were performed in all neonates. Brain injury was defined by the presence of intraventricular hemorrhage grade III/IV, cystic periventricular leukomalacia or infarct. Peripheral blood samples were collected from all neonates on days(d) 1, 3, 9, 18 and 45 of life for the measurement of levels of CPCs [early and late Endothelial Progenitor Cells (EPCs), Haematopoietic Stem Cells (HSCs) and Very Small Embryonic-Like Stem Cells (VSELs)], Neuron-Specific Enolase (NSE), S100b, Erythropoietin (EPO) and Stromal Cell-Derived Factor-1 (SDF-1) . RESULTS: Ten out of the 23 preterm infants included in the study developed brain injury; the remaining thirteen infants served as controls. In the brain injury group a significant increase of HSCs (d9, d45), early EPCs (d3, d9, d18) and late EPCs (d1, d3, d9, d18, d45) was observed compared to controls. VSELs on d45 were significantly higher in controls. S100b on d1, EPO on d1, SDF-1 on d3 and NSE on d18 were significantly increased in the brain injury group. Moreover, CPCs were significantly related to S100b, NSE, EPO and SDF-1 levels at multiple time points. CONCLUSIONS: The observed pattern of CPCs mobilization and its association with biomarkers following brain injury in preterm neonates indicate the existence of an endogenous brain regeneration process. Enhancement of this process with exogenous progenitor cell transplantation might be a powerful therapeutic strategy to restore brain damage and improve the neurodevelopmental outcome in premature infants. Hippokratia 2015; 19 (2):141-147.

Intratumoral gene therapy versus intravenous gene therapy for distant metastasis control with 2-diethylaminoethyl-dextran methyl methacrylate copolymer non-viral vector-p53.

Lung cancer still remains to be challenged by novel treatment modalities. Novel locally targeted routes of administration are a methodology to enhance treatment and reduce side effects. Intratumoral gene therapy is a method for local treatment and could be used either in early-stage lung cancer before surgery or at advanced stages as palliative care. Novel non-viral vectors are also in demand for efficient gene transfection to target local cancer tissue and at the same time protect the normal tissue. In the current study, C57BL/6 mice were divided into three groups: (a) control, (b) intravenous and (c) intatumoral gene therapy. The novel 2-Diethylaminoethyl-Dextran Methyl Methacrylate Copolymer Non-Viral Vector (Ryujyu Science Corporation) was conjugated with plasmid pSicop53 from the company Addgene for the first time. The aim of the study was to evaluate the safety and efficacy of targeted gene therapy in a Lewis lung cancer model. Indeed, although the pharmacokinetics of the different administration modalities differs, the intratumoral administration presented increased survival and decreased distant metastasis. Intratumoral gene therapy could be considered as an efficient local therapy for lung cancer.

Intramyocardial implantation of autologous bone marrow-derived stem cells combined with coronary artery bypass grafting in patients with ischemic cardiomyopathy: a pilot study.

BACKGROUND: Ischemic cardiomyopathy has the distinctiveness of irreversible myocardial damage with scar tissue formation and mainly impaired perfusion of the remaining viable myocardium. We present results of the first series of patients with severe ischemic cardiomyopathy managed in our institution with intramyocardial implantation of autologous bone marrow stem cells at the time of coronary artery bypass grafting. The aim is to evaluate feasibility and safety of the procedure in our institution. PATIENTS AND METHODS: Nine patients with severe ischemic cardiomyopathy scheduled for elective coronary artery bypass grafting were managed with concurrent intramyocardial autologous bone marrow stem cells injection in pre-defined viable peri-infarct areas that showed poor perfusion and could not be grafted. Detailed mapping of infracted and hibernating myocardial segments was performed in all patients with single photon emission computed tomography segmental analysis. RESULTS: There was no perioperative 30-day mortality. Improvement was evident in left ventricular ejection fraction which was increased significantly from 31.3% preoperatively to 42.4%, 46.6% and 52.5% at 3, 6 and 12 months respectively. Postoperative thallium scintigraphy revealed increased perfusion in myocardial segments corresponding to areas of stem cell injection and a net reduction in the estimated infarct size at 6 and 12 months in 5/8 (62.5%) patients. CONCLUSIONS: Preliminary data from this pilot study show that intramyocardial administration of bone marrow stem cells in patients undergoing coronary bypass grafting for ischemic cardiomyopathy is safe and associated with an improvement in left ventricular function and enhanced reperfusion of non-viable myocardial territories.

The effects of tacrolimus on colonic anastomotic healing in rats.

AIM: The aim of this experimental study is to investigate the effects of tacrolimus on colonic anastomotic healing after subcutaneous administration. MATERIALS AND METHODS: Forty Albino-Wistar male rats were divided into two groups, with two equal subgroups each. They all underwent colonic resection followed by a single-layer, inverted colon anastomosis and were injected subcutaneously with either 1 ml of 0.9% NaCl solution or tacrolimus (0.1 mg/kg body weight) depending on their group. Half of the rats were sacrificed on the fourth postoperative day, while the remaining half were sacrificed on the eighth postoperative day. Macroscopical and histological assessment was performed, while anastomotic bursting pressures and the tissue concentrations in hydroxyproline and collagenase I were evaluated. RESULTS: On the fourth postoperative day, the bursting pressures (217.00 ± 11.12, p < 0.001), the fibroblast activity (2.80 ± 0.42, p = 0.022), the neoangiogenesis (2.10 ± 0.32, p = 0.007) and the tissue hydroxyproline concentration (254.23 ± 67.10, p = 0.001) were significantly higher in the tacrolimus-treated animals. Furthermore, tacrolimus significantly decreased the inflammatory cell infiltration (1.50 ± 0.53, p < 0.001) and the tissue collagenase I concentration (4.16 ± 0.76, p = 0.002). On the eighth day, the bursting pressure (264.00 ± 32.61, p < 0.001) and the hydroxyproline tissue concentration (331.04 ± 55.56, p = 0.002) were significantly higher in the tacrolimus subgroups. The inflammatory cell infiltration (1.20 ± 0.42, p < 0.001) and the collagenase I concentration (1.61 ± 0.83, p < 0.001) were significantly lower. In addition, the adhesion formation score was significantly lower (1.20 ± 0.92, p = 0.065). CONCLUSION: Tacrolimus, when injected subcutaneously, promotes healing of colonic anastomoses in rats. It impairs not only inflammatory response but also collagen degradation, resulting to increased anastomotic strength on the fourth as well as on the eighth postoperative day.

Intraperitoneally administered irinotecan with 5-fluorouracil impair wound healing of colonic anastomoses in a rat model: an experimental study.

AIM: The aim of this experimental study is the assessment of the effects of the immediate post-operative intraperitoneal administration of 5-fluorouracil and irinotecan on the healing process of large bowel anastomoses in rats. MATERIALS AND METHODS: Sixty male Wistar rats were divided into 4 groups of 15 rats each. The rats underwent large bowel resection and anastomosis, followed by the intraperitoneal administration of normal saline (group 1), 5-fluorouracil (group 2), irinotecan (group 3) or the combination of 5-fluorouracil and irinotecan (group 4). All animals were killed on the eighth post-operative day. During post-mortem examination, the anastomoses were assessed macroscopically for a possible anastomotic leak and the extent of adhesion formation. Subsequently, the anastomotic bursting pressure was measured, and the anastomoses were assessed histologically. RESULTS: No anastomotic dehiscence was observed in the rats of group 1. In groups 2 and 3, we observed 3 anastomotic leaks in each group, and in group 4, we observed 5 leaks (P = 0.111). The mean bursting pressure of the anastomoses in group 1 was significantly higher compared to groups 2, 3 and 4 (P < 0.001). The least inflammatory cell infiltration score was observed in group 1 (P < 0.001). The lowest neoangiogenesis score was observed in group 2 and the highest in group 4. The collagen formation in group 1 was significantly higher compared to the other 3 groups (P < 0.001). Similar results were observed for the fibroblast activity, where group 1 revealed significantly higher fibroblast scores compared to groups 2, 3 and 4 (P < 0.001). Finally, groups 2, 3 and 4 showed significantly lower hydroxyproline levels compared to the control group (P < 0.001). CONCLUSION: The immediate, post-operative intraperitoneal administration of 5-fluorouracil or irinotecan had a negative effect on the healing process of the large bowel anastomoses in rats. The negative effects of the combination of 5-fluorouracil and irinotecan were statistically more significant compared to the single use of 5-fluorouracil or irinotecan.

The effects of iloprost on colonic anastomotic healing in rats.

PURPOSE: The purpose of this experimental study is to investigate the effects of iloprost on colonic anastomotic healing in rats, after intraperitoneal administration. METHODS: Forty male Albino-Wistar rats were randomized into two groups of twenty animals each. They all underwent colonic resection followed by an inverted anastomosis. The rats of Group A (control) received 3 ml of NaCl intraperitoneally, while those of Group B (iloprost) received iloprost (2 µg/kg body weight), immediately postoperatively and daily until killed. Each group was further divided into two equal subgroups, depending on the day of killing. The animals of subgroups 1 were killed on the fourth postoperative day, while those of subgroups 2 on the eighth. Macroscopical and histological assessments were performed. Besides, anastomotic bursting pressures and the tissue concentrations in hydroxyproline and collagenase I were also evaluated. RESULTS: No anastomotic dehiscence was noted. The mean bursting pressure was higher in the iloprost group compared with the control group, but a significant difference was revealed only on the fourth postoperative day. Furthermore, iloprost significantly increased the new vessel formation on the fourth, as well as on the eighth postoperative day. CONCLUSION: Iloprost enhances the early phase of colonic anastomotic healing in rats.

The effects of the intraperitoneal administration of oxaliplatin and 5-FU on the healing of colonic anastomoses: an experimental study.

BACKGROUND: The purpose of this experimental study was to assess the effects of the immediate postoperative intraperitoneal administration of oxaliplatin and 5-FU on the healing of colonic anastomoses in rats. METHODS: Sixty rats were randomized into 4 groups of 15 rats each and were subjected to colonic anastomoses. To the 1st group, saline solution was administered immediately postoperatively, intraperitoneally. To the 2nd group, 5-FU was administered, to the 3rd group oxaliplatin and to the 4th group 5-FU and oxaliplatin were administered immediately postoperatively, intraperitoneally. After killing the rats on the 8th postoperative day, the anastomoses were examined macroscopically and the anastomotic bursting pressures were measured. The anastomoses were also examined histologically and the hydroxyproline contents were determined. RESULTS: Rupture of the anastomosis was observed in no rats of the 1st group, in 3 rats of the 2nd group, in 4 rats of the 3rd group and in 7 rats of the 4th group (P = 0.016). The bursting pressure (P < 0.001), the hydroxyproline content (P < 0.001) and the concentration of collagen (P < 0.001) and fibroblasts (P < 0.001) were significantly lower in the 2nd, 3rd and 4th group in comparison with the 1st group. The formation of adhesions and the leukocytosis on the anastomoses were significantly higher in the 2nd, 3rd and 4th group than in the 1st group (P < 0.001). CONCLUSIONS: The immediate postoperative, intraperitoneal administration of oxaliplatin, 5-FU or the combination of 5-FU and oxaliplatin impairs the healing of colonic anastomoses in rats.

Isolation of mesenchymal stem cells using the total length of umbilical cord for transplantation purposes.

BACKGROUND: Umbilical cord (UC) mesenchymal cells have the ability to differentiate into various cell types, which make them an easily obtainable source for therapeutic uses. Different approaches have been used to isolate mesenchymal stem cells (MSC). AIM: Here, we report a detailed enzymatic method where large number of cells can be efficiently isolated from the cord matrix and cryopreserved on the same day of arrival at the laboratory. METHODS/MATERIALS: Cells were successfully isolated from 12 samples, with a new procedure that uses the total length of the UC. MSC have been isolated using a detailed enzymatic method with collagenase and hyaluronidase followed by trypsin, without removing the vessels and without mincing the cord. Stem cells were measured with flow cytometry before cryopreservation and post-thaw. Cultured cells were assessed for MSC marker expression and adherence plasticity for three passages. Multilineage differentiation was performed. RESULTS: Nucleated cell yield was calculated at 0·95 × 10(6) /cm. MSC yield was calculated at 0·65 × 10(6) /cm of cord with flow cytometry while the mean length was 31 cm. Cultured cells expressed the mesenchymal markers CD29, CD90, CD105 and CD44. Mesenchymal marker expression remained intact over the three passages and post-thaw. Osteogenic and adipogenic differentiation was evaluated. CONCLUSIONS: Our findings provide a fast and efficient method for mesenchymal cell isolation from Wharton's jelly using the total length of the UC. This method resulted in a large number of cells while the cells retained their mesenchymal character after thawing. This method can be easily applied, along with UC blood, for UC banking.

Intraperitoneal administration of bevacizumab intraoperatively does not affect abdominal wound healing in rats.

BACKGROUND: Bevacizumab is a monoclonal antibody targeted at vascular endothelial growth factor (VEGF) to treat advanced colorectal cancer as well as other malignancies, but the ideal time point for its administration in patients scheduled for surgery is not well defined due to serious concerns regarding possible side effects on wound healing. Therefore, we conducted an experimental study in rats to clarify this issue. METHODS: Four groups of 10 Wistar rats each underwent a 4-cm midline laparotomy and closure of the wound in 2 layers. In the treatment groups (A and B), bevacizumab (Avastin(®)) received a single dose of 5 mg/kg i.p., and an equal amount of saline was given to the control groups (C and D). Groups A and C were sacrificed on the 7th postoperative day, and groups B and D on the 14th postoperative day. Wounds were inspected by two independent observers upon sacrifice and results were recorded; wound tissues were sent for histology to assess the degree of fibrosis and measurement of tissue hydroxyproline levels. Serum levels of endothelin-1, C-reactive protein, pro-oxidant/antioxidant balance and carbonylated proteins were also determined. For statistical analysis, the Mann-Whitney U test was used. RESULTS: Wound healing did not differ among groups both on the 7th and the 14th postoperative days, and there was also no significant difference regarding the degree of inflammation, fibroblast proliferation and collagen synthesis, as well as hydroxyproline and biochemical marker levels among the groups. CONCLUSIONS: Intraperitoneal bevacizumab administered intraoperatively does not significantly affect abdominal wound healing in rats.

Combination of P300 and CSF ß-amyloid(1-42) assays may provide a potential tool in the early diagnosis of Alzheimer's disease.

BACKGROUND/AIMS: The aim of this study was to investigate the diagnostic role of CSF beta amyloid(1-42) levels and auditory event-related potentials (AERPs) in the progression of mild cognitive impairment (MCI) to Alzheimer's disease (AD). METHODS: In fifty three MCI patients a lumbar puncture was performed and beta amyloid(1-42) levels were determined. Twenty patients were re-examined after 11 months. During this period five of them progressed to AD. Neuropsychological and ERP examinations were performed in all patients at both exams. RESULTS: Compared to MCI stable patients, AD-converters showed significantly lower beta-amyloid(1-42) values both for group 1 (Mann Whitney test, Z=-2.952, p=0.003, effect size r=-0.41) and group 2 (Z=-2.458, p=0.011; effect size r=-0.55). On the other hand, the patients of group 1 who converted to AD had prolonged latencies and lower amplitudes of the P300 wave compared to those of the MCI-stable patients, although the differences were not significant. CONCLUSIONS: Compared to the separate use of CSF beta-amyloid(1-42) and AERPs, higher values of sensitivity and specificity were achieved by the combined use of beta-amyloid(1-42) levels and P300 latencies (80% and 98%) or amplitudes (100% and 89%) in the discrimination between AD converters and MCI stable patients. Therefore the combination of an electrophysiological and a biological marker is potentially of high diagnostic value for the early diagnosis of AD converters.

Addition of adipose-derived stem cells in cord blood cultures stimulates their pluripotent differentiation.

INTRODUCTION: Adipose tissue is recognized as an important source of postnatal mesenchymal stem cells for generative medicine applications. Moreover, cord blood stem cells have been shown to contain pluripotent stem cells called unrestricted somatic stem cells (USSCs). However, this population is rare and cannot be generated from every cord blood sample. In this study, we have presented a new method of co-culture of adipose-derived stem cells (ADPCs) and cord blood stem cells that results in pluripotent differentiation. MATERIALS AND METHODS: ADPCs were obtained from a piece of adipose tissue after treatment with 0.075% collagenase, which was subsequently inactivated with DMEM/10% FBS. The cellular pellet of centrifugation was plated at 5-7 x 10(6) cells/mL in T25 culture flasks in a low-glycose DMEM with 30% FCS. Cord blood stem cells were obtained by centrifugation following double-processing in the presence of 2% HES 200/0.5 and plated at 5-7 x 10(6) cells/mL in the same medium. To investigate the crucial role of ADPCs in pluripotent cord blood differentiation, we added a ADPCS as (1 x 10(4) cells/mL) to the cord blood cultures and analyzed the contribution of ADPCs using a microscope as well as with flow cytometry. RESULTS: After only 3 days, adherent cells (USSC colonies) of fibroblastic morphology were detected in all co-cultured samples, whereas this was observed later or not at all in the non-co-cultured samples. The greater density of colonies in the co-coltured samples was another point. Hematopoietic CD45 cells were no longer detected after the first passage. Pluripotent stem cells were obtained from all co-cultured samples that contained stem cells positive for CD29, CD44, CD49e, CD90, CD105, CD51 Stro, and C-kit antibodies but negative for CD34, CD45, CD133, and glycophorin A. CONCLUSION: Addition of ADPCs was crucial to generate pluripotent-derived stem cells from cord blood samples. This double culture may be a useful tool for a universal allogeneic stem cell source for tissue repair or regeneration.

Progression of mild cognitive impairment to Alzheimer's disease: improved diagnostic value of the combined use of N200 latency and beta-amyloid(1-42) levels.

BACKGROUND/AIMS: The aim of this study was to investigate the role of cerebrospinal fluid beta-amyloid(1-42) levels and auditory event-related potentials (AERPs) in the progress of mild cognitive impairment (MCI) to Alzheimer's disease (AD). METHODS: In 53 MCI patients, lumbar puncture was performed and beta-amyloid(1-42) levels were determined. Twenty patients were reexamined after 11 months. During this period, 5 of them progressed to AD. Neuropsychological and ERP analyses were performed on all patients during both baseline and endpoint examinations. RESULTS: Compared to stable MCI patients, those that progressed to AD had significantly lower beta-amyloid(1-42) levels (Mann-Whitney test, Z = -2.952, p = 0.003; effect size r = -0.41) and significantly prolonged N200 latencies (Mann-Whitney test, Z = -3.561, p < 0.001, effect size r = -0.49). From ERP variables, only the N200 latency significantly correlated with beta-amyloid(1-42) levels (baseline examination: r(s) = -0.421, p = 0.002; follow-up examination: r(s) = -0.574, p = 0.008). CONCLUSIONS: The combined use of these two parameters enabled discrimination of stable MCI patients from those who developed AD, with 100% sensitivity and specificity. Therefore, this method could be of high diagnostic value for the early diagnosis of AD.

Prediction of conversion from mild cognitive impairment to Alzheimer's disease by CSF cytochrome c levels and N200 latency.

The aim of the present study was to investigate the role of CSF cytochrome c levels and auditory event-related potentials (AERPs) on the progress of mild cognitive impairment (MCI) to Alzheimer's disease (AD). Fifty one patients diagnosed with MCI and fourteen healthy individuals underwent lumbar puncture at baseline and their CSF cytochrome c levels were determined. A follow-up examination of cytochrome c levels took place in 20 patients after 11 months and in this period five of the patients progressed to AD. ERP examinations were also performed in all patients both at baseline and follow-up. MCI patients had significantly higher cytochrome c levels compared to healthy controls (Mann-Whitney test, Z=-2.110, p=0.018). Compared to MCI patients who remained stable, the AD-converters, had a higher increase over time in cytochrome c levels (Mann-Whitney test, p=0.002; effect size r=0.63) and significantly prolonged N200 latency (Mann-Whitney test, p<0.001; effect size r=0.50). Amongst investigated ERP variables, only N200 amplitude was significantly correlated with CSF cytochrome c levels (rs=0.310, p=0.03). Both parameters were proved capable of discriminating AD converters from those MCI patients who remained stable, with sensitivity and specificity >75%. Our results suggest that conversion from MCI to AD is associated with a marked elevated N200 latency at baseline and a high increase in cytochrome c levels during a relatively short period of time, and that both parameters could be possibly considered as candidate markers for the discrimination between MCI patients who convert to AD and those who remain stable.

Late postoperative opacification of hydrogel intraocular lenses: analysis of 13 explanted lenses.

PURPOSE: We report the clinical, morphological, and ultrastructural findings of 13 consecutively explanted opacified Hydroview(R) (hydrogel) intraocular lenses (IOLs). Our purpose was to provide a comprehensive account on the possible factors involved in late postoperative opacification of these IOLs. PATIENTS AND METHODS: Thirteen consecutive opacified hydrogel IOLs (Hydroview H 60 M, Bausch & Lomb) were explanted due to the significant visual impairment they caused. The IOLs underwent macroscopical examination, transmission electron microscopy (TEM), scanning electron microscopy (SEM), energy-dispersive X-ray spectroscopy (EDS), and electrophoresis for protein detection. Three unused control Hydroview IOLs served for comparison. RESULTS: Macroscopical examination showed a diffuse or localized grey-whitish opacification within the IOL optic. TEM confirmed the presence of lesions inside the optic in all the explanted IOLs and revealed 3 patterns of deep deposits: a) diffuse, thick, granular, electron-dense ones; b) small, thin, lattice-like ones, with prominent electron-lucent areas; and c) elongated electron-dense formations surrounded by electron-lucent halos. SEM showed surface deposits on four IOLs. EDS revealed oxygen and carbon in all IOLs and documented calcium, phosphorus, silicon and/or iron in the deposits. Two of the patients with iron in their IOLs had eye surgery prior to their phacoemulsification. Iron correlated well with the second TEM pattern of deep lesions, whereas calcium with the third TEM pattern. No protein bands were detected on electrophoresis. Control lenses did not show any ultrastructural or chemical abnormality. CONCLUSIONS: The present study supports the presence of chemical alterations inside the polymer of the optic in late postoperative opacification of Hydroview IOLs. This opacification does not follow a unique pathway but may present under different ultrastructular patterns depending on the responsible factors. Mechanical stress during surgery may initiate a sequence of events where ions such as calcium, phosphorus, silicon, and/or iron, participate in a biochemical cascade that leads to gradual alteration of the polymer network. Intraocular inflammation due to previous operation may be a factor inducing opacification through increase of iron-binding capacity in the aqueous humour. Calcification accounts only partially for the opacification noted in this type of IOL.

Oxidative effects of inorganic and organic contaminants on haemolymph of mussels.

We applied a newly-established method in haemolymph of mussels, Mytilus galloprovincialis, exposed to different concentrations of heavy metals, such as zinc and cadmium and organic pollutants, such as PAHs and lindane, for the detection of total antioxidant capacity (TAC). The susceptibility of exposed mussels was increased in relation to oxidative stress induced by contaminants tested. Oxidative modifications of proteins were estimated by measuring protein carbonyl content (PCC) and malondialdehyde levels (MDA). For PCC measurement, a highly sensitive and accurate ELISA method, which requires only 5 microg of protein, was used. The significant increase of PCC and MDA in haemolymph of exposed mussels reinforces its role as biomarkers of oxidative stress. Significant correlation of TAC assay, PCC and MDA was conducted in order to evaluate the utility of PCC and TAC assay, used in the present study, as tools for determining oxidative effects of pollutants in mussels. The results reinforce the application of PCC method as useful tool for the determination of PCC alterations in haemolymph of mussels exposed to different levels of contaminants. In addition, the TAC method gives encouraging results, concerning its ability to predict antioxidant efficiency in haemolymph of mussels exposed to inorganic and organic contaminants.

Insulin sensitivity increase after calcium supplementation and change in intraplatelet calcium and sodium-hydrogen exchange in hypertensive patients with Type 2 diabetes.

AIMS/HYPOTHESIS: To investigate the effect of oral calcium (Ca(2+)) supplementation on insulin sensitivity measured by the euglycaemic hyperinsulinaemic clamp, intraplatelet cationic concentration of Ca(2+) ([Ca(2+)](i)) and the transmembrane sodium-hydrogen exchanger (NHE) activity in erythrocytes in subjects with Type 2 diabetes and hypertension. PATIENTS AND METHODS: In this parallel randomized controlled single-blinded trial, 31 patients were allocated to receive either 1500 mg of Ca(2+) orally, daily (n = 15) or no treatment (n = 16) for 8 weeks. At baseline and at the end of the 8-week period insulin sensitivity, [Ca(2+)](i) and the first isoform of NHE (NHE-1) activity were measured. RESULTS: At the end of the study, subjects who received Ca(2+) supplementation showed higher insulin sensitivity (Delta M-value 0.32 +/- 0.5 mmol/min P < 0.05) and lower [Ca(2+)](i) (125.0 +/- 24.7 to 80.4 +/- 10.6 nmol/l, P < 0.05, mean +/- sem) and NHE-1 activity (79.5 +/- 10.0 to 52.1 +/- 6.4 mmol Na/l red cell/h, P < 0.05). None of the above parameters were changed in the control group. Simple regression analysis demonstrated the change in [Ca(2+)](i) significantly determined insulin sensitivity change (beta = -0.36, P < 0.05). CONCLUSIONS/INTERPRETATION: Oral Ca(2+) supplementation improves insulin sensitivity in patients with Type 2 diabetes and hypertension. These changes are likely to be mediated by changes in intracellular ionic Ca(2+). NHE-1 activity was also reduced after Ca(2+) supplementation but its role in insulin sensitivity requires further investigation.