Evaluation of Polysaccharide-Peptide Conjugates Containing the RGD Motif for Potential Use in Muscle Tissue Regeneration.

New scaffold materials composed of biodegradable components are of great interest in regenerative medicine. These materials should be: stable, nontoxic, and biodegrade slowly and steadily, allowing the stable release of biodegradable and biologically active substances. We analyzed peptide-polysaccharide conjugates derived from peptides containing RGD motif (H-RGDS-OH (1), H-GRGDS-NH2 (2), and cyclo(RGDfC) (3)) and polysaccharides as scaffolds to select the most appropriate biomaterials for application in regenerative medicine. Based on the results of MTT and Ki-67 assays, we can state that the conjugates containing calcium alginate and the ternary nonwoven material were the most supportive of muscle tissue regeneration. Scanning electron microscopy imaging and light microscopy studies with hematoxylin-eosin staining showed that C2C12 cells were able to interact with the tested peptide-polysaccharide conjugates. The release factor (Q) varied depending on both the peptide and the structure of the polysaccharide matrix. LDH, Alamarblue®, Ki-67, and cell cycle assays indicated that peptides 1 and 2 were characterized by the best biological properties. Conjugates containing chitosan and the ternary polysaccharide nonwoven with peptide 1 exhibited very high antibacterial activity against Staphylococcus aureus and Klebsiella pneumoniae. Overall, the results of the study suggested that polysaccharide conjugates with peptides 1 and 2 can be potentially used in regenerative medicine.

Screening of Self-Assembling of Collagen IV Fragments into Stable Structures Potentially Useful in Regenerative Medicine.

The aim of the research was to check whether it is possible to use fragments of type IV collagen to obtain, as a result of self-assembling, stable spatial structures that could be used to prepare new materials useful in regenerative medicine. Collagen IV fragments were obtained by using DMT/NMM/TosO- as a coupling reagent. The ability to self-organize and form stable spatial structures was tested by the CD method and microscopic techniques. Biological studies covered: resazurin assay (cytotoxicity assessment) on BJ, BJ-5TA and C2C12 cell lines; an alkaline version of the comet assay (genotoxicity), Biolegend Legendplex human inflammation panel 1 assay (SC cell lines, assessment of the inflammation activity) and MTT test to determine the cytotoxicity of the porous materials based on collagen IV fragments. It was found that out of the pool of 37 fragments (peptides 1-33 and 2.1-2.4) reconstructing the outer sphere of collagen IV, nine fragments (peptides: 2, 4, 5, 6, 14, 15, 25, 26 and 30), as a result of self-assembling, form structures mimicking the structure of the triple helix of native collagens. The stability of spatial structures formed as a result of self-organization at temperatures of 4 °C, 20 °C, and 40 °C was found. The application of the MST method allowed us to determine the Kd of binding of selected fragments of collagen IV to ITGα1ß1. The stability of the spatial structures of selected peptides made it possible to obtain porous materials based on their equimolar mixture. The formation of the porous materials was found for cross-linked structures and the material stabilized only by weak interactions. All tested peptides are non-cytotoxic against all tested cell lines. Selected peptides also showed no genotoxicity and no induction of immune system responses. Research on the use of porous materials based on fragments of type IV collagen, able to form stable spatial structures as scaffolds useful in regenerative medicine, will be continued.

Conjugates of Copper Alginate with Arginine-Glycine-Aspartic Acid (RGD) for Potential Use in Regenerative Medicine.

Current restrictions on the use of antibiotics, associated with increases in bacterial resistance, require new solutions, including materials with antibacterial properties. In this study, copper alginate fibers obtained using the classic wet method were used to make nonwovens which were modified with arginine-glycine-aspartic acid (RGD) derivatives. Stable polysaccharide-peptide conjugates formed by coupling with 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)-4-methylmorpholinium toluene-4-sulfonate (DMT/NMM/TosO-), and materials with physically embedded RGD derivatives, were obtained. The materials were found to be characterized by very high antibacterial activity against S. aureus and K. pneumoniae. Cytotoxicity studies confirmed that the materials are not cytotoxic. Copper alginate conjugates with RGD peptides have strong potential for use in regenerative medicine, due to their biocompatibility and innate antibacterial activity.

VASIM: an automated tool for the quantification of carotid atherosclerosis by computed tomography angiography.

The diagnostic imaging techniques currently used to evaluate the arterial atherosclerosis hinge on the manual marking and calculation of the stenosis degree. However, the manual assessment is highly dependent on the operator and characterized by low replicability. The study aimed to develop a fully-automated tool for the segmentation and analysis of atherosclerosis in the extracranial carotid arteries. The dataset consisted of 59 randomly-chosen individuals who had undergone head-and-neck computed tomography angiography (CTA), at the Tampere University Hospital, Tampere, Finland. The analysis algorithm was mainly based on the detection of carotid arteries, delineation of the vascular wall, and extraction of the atherosclerotic plaque. To improve the vascular detection rate, the model-based and volume-wide analytical approaches were deployed. A new fully-automated vascular imaging (VASIM) software tool was developed. For stenosis over 50%, the success rate was 83% for the detection and segmentation. Specificity and sensitivity of the algorithm were 25% and 83%, respectively. The overall accuracy was 71%. The VASIM tool is the first published approach for the fully-automated analysis of atherosclerosis in extracranial carotid arteries. The tool provides new outputs, which may help with the quantitative and qualitative, clinical evaluation of the atherosclerosis burden and evolution. The findings from this study provide a basis for the further development of automated atherosclerosis diagnosis and plaque analysis with CTA.

Diffusion tensor imaging and disability progression in multiple sclerosis: A 4-year follow-up study.

OBJECTIVES: Diffusion tensor imaging (DTI) is sensitive technique to detect widespread changes in water diffusivity in the normal-appearing white matter (NAWM) that appears unaffected in conventional magnetic resonance imaging. We aimed to investigate the prognostic value and stability of DTI indices in the NAWM of the brain in an assessment of disability progression in patients with a relapsing-onset multiple sclerosis (MS). METHODS: Forty-six MS patients were studied for DTI indices (fractional anisotropy (FA), mean diffusivity (MD), radial (RD), and axial (AD) diffusivity) in the NAWM of the corpus callosum (CC) and the internal capsule at baseline and at 1 year after. DTI analysis for 10 healthy controls was also performed at baseline. Simultaneously, focal brain lesion volume and atrophy measurements were done at baseline for MS patients. Associations between DTI indices, volumetric measurements, and disability progression over 4 years were studied by multivariate logistic regression analysis. RESULTS: At baseline, most DTI metrics differed significantly between MS patients and healthy controls. There was tendency for associations between baseline DTI indices in the CC and disability progression (p < 0.05). Changes in DTI indices over 1 year were observed only in the CC (p < 0.008), and those changes were not found to predict clinical worsening over 4 years. Clear-cut association with disability progression was not detected for baseline volumetric measurements. CONCLUSION: Aberrant diffusivity measures in the NAWM of the CC may provide additional information for individual disability progression over 4 years in MS with the relapsing-onset disease. CC may be a good target for DTI measurements in monitoring disease activity in MS, and more studies are needed to assess the related prognostic potential.

SMAD-4 gene expression in human colorectal cancer: Comparison with some clinical and pathological parameters.

The aim of this study was to evaluate the expression of SMAD-4 gene in cases of colorectal cancer and to link the obtained data with the development of this disease. SMAD-4 gene is responsible for the control of many important cellular processes, for example prevention of excessive epithelial cell growth and divisions. This suppressor gene is located on chromosome 18 within the region with frequent genetic losses in colorectal cancer. Inactivation of this gene is commonly found in pancreatic cancer where the SMAD-4 gene lost in the expression has been associated with a poor prognosis in this cancer. However, the role of SMAD-4 gene in other cancers has not been completely explained, therefore in the present study we tried to find the role of this gene in colon cancer. The relative expression level of SMAD-4 gene was determined by real-time PCR for 80 cases of colorectal cancer. The obtained results for SMAD-4 expression were compared with many clinical and pathological variables (such as the size and depth of primary tumour penetration, presence of the metastases, stage of cancer, histological grade or overall survival). It was found that the level of SMAD-4 gene expression was not associated with the analyzed parameters of clinical staging. The lack of dependence can be caused by slight differences within the study group in view of parameters correlated with invasive colon cancer. Further analysis in this direction is needed.

Association between soluble L-selectin and anti-JCV antibodies in natalizumab-treated relapsing-remitting MS patients.

OBJECTIVE: In relapsing-remitting MS (RRMS) patients treated with natalizumab, the low level of L-selectin-expressing CD4+ T cells has been associated with the risk of progressive multifocal leukoencephalopathy (PML). In this study, our aim was to correlate the levels of soluble L-selectin and the anti-JCV antibody index in the sera of RRMS patients treated with natalizumab. METHODS: This study included 99 subjects, including 44 RRMS patients treated with natalizumab, 30 with interferon beta (IFN-ß) and 25 healthy controls. The levels of soluble L-selectin (sL-selectin) in sera were measured by ELISA, and the anti-JC Virus (JCV) antibody index was determined by the second-generation ELISA (STRATIFY JCV™ DxSelect™) assay. RESULTS: A significant correlation was found between the levels of sL-selectin and anti-JCV antibody indices in sera in the natalizumab-treated patients (r=0.402; p=0.007; n=44), but not in those treated with IFN-ß. This correlation became even stronger in JCV seropositive patients treated with natalizumab for longer than 18 months (r=0.529; p=0.043; n=15). CONCLUSION: The results support the hypothesis of sL-selectin being connected to the anti-JCV antibody index values and possibly cellular L-selectin. Measurement of serum sL-selectin should be evaluated further as a potential biomarker for predicting the risk of developing PML.

Analysis of apoptosis-related genes in patients with clinically isolated syndrome and their association with conversion to multiple sclerosis.

To analyse whether the expression of apoptotic transcripts is associated with the conversion from clinically isolated syndrome (CIS) to multiple sclerosis (MS). Eleven candidate transcripts belonging to the death receptor pathway, BCL-2, the inflammasome complex and NF-ΚB family were studied in the nonconverting and converting CIS patients during the four-year follow-up period. Conversion to MS was associated with marked variability in the expression of proapoptotic genes that were linked to TGF-B1 gene levels. The predominant expression of proapoptotic genes in patients with CIS suggests an increased potential to undergo apoptosis with the goal of terminating immune responses and regulating immune system homeostasis.

Longitudinal assessment of clinically isolated syndrome with diffusion tensor imaging and volumetric MRI.

The potential of diffusion tensor imaging (DTI) indices and volumes of focal lesions on conventional magnetic resonance imaging to predict conversion to multiple sclerosis (MS) was analyzed in subjects with clinically isolated syndrome (CIS) over 4 years. Twenty patients with CIS and 10 healthy controls were included in the study. The data showed an association between the volumes of T1 and fluid-attenuated inversion recovery (FLAIR) lesions and conversion to MS (T1: P=.02; FLAIR: P=.02). The worsening of DTI indices (mean diffusivity and fractional anisotropy) was primarily seen in patients progressing to MS, but clear-cut association with conversion could not be detected.

[Primary testical lymphoma]. / Pierwotny chloniak jadra.

In our report we present a rare case of primary non Hopdgkin lymphoma of the testis in 70-years-old man, who was diagnosed and treated from syderopenic anemia. Here we demonstrate difficulties with differential diagnosis mentioned tumor.