Macrophage cell morphology-imprinted substrates can modulate mesenchymal stem cell behaviors and macrophage M1/M2 polarization for wound healing applications

Mesenchymal stem cells and macrophages (MQ) are two very important cells involved in the normal wound healing process. It is well understood that topological cues and mechanical factors can lead to different responses in stem cells and MQ by influencing their shape, cytoskeleton proliferation, migration, and differentiation, which play an essential role in the success or failure of biomaterial implantation and more importantly wound healing. On the other hand, the polarization of MQ from proinflammatory (M1) to prohealing (M2) phenotypes has a critical role in the acceleration of wound healing. In this study, the morphology of different MQ subtypes (M0, M1, and M2) was imprinted on a silicon surface (polydimethylsiloxane [PDMS]) to prepare a nano-topography cell-imprinted substrate with the ability to induce anti-inflammatory effects on the mouse adipose-derived stem cells (ADSCs) and RAW264.7 monocyte cell line (MO). The gene expression profiles and flow cytometry of MQ revealed that the cell shape microstructure promoted the MQ phenotypes according to the specific shape of each pattern. The ELISA results were in agreement with the gene expression profiles. The ADSCs on the patterned PDMS exhibited remarkably different shapes from no-patterned PDMS. The MOs grown on M2 morphological patterns showed a significant increase in expression and section of anti-inflammatory cytokine compared with M0 and M1 patterns. The ADSCs homing in niches heavily deformed the cytoskeletal, which is probably why the gene expression and phenotype unexpectedly changed. In conclusion, wound dressings with M2 cell morphology-induced surfaces are suggested as excellent anti-inflammatory and antiscarring dressings.     

Control of Blood Pressure and Risk Attenuation: Post Trial Follow-Up of Randomized Groups

Background

Evidence on long term effectiveness of public health strategies for lowering blood pressure (BP) is scarce. In the Control of Blood Pressure and Risk Attenuation (COBRA) Trial, a 2 x 2 factorial, cluster randomized controlled trial, the combined home health education (HHE) and trained general practitioner (GP) intervention delivered over 2 years was more effective than no intervention (usual care) in lowering systolic BP among adults with hypertension in urban Pakistan. However, it was not clear whether the effect would be sustained after the cessation of intervention. We conducted 7 years follow-up inclusive of 5 years of post intervention period of COBRA trial participants to assess the effectiveness of the interventions on BP during extended follow-up.

Methods

A total of 1341 individuals 40 years or older with hypertension (systolic BP 140 mm Hg or greater, diastolic BP 90 mm Hg or greater, or already receiving treatment) were followed by trained research staff masked to randomization status. BP was measured thrice with a calibrated automated device (Omron HEM-737 IntelliSense) in the sitting position after 5 minutes of rest. BP measurements were repeated after two weeks. Generalized estimating equations (GEE) were used to analyze the primary outcome of change in systolic BP from baseline to 7- year follow-up. The multivariable model was adjusted for clustering, age at baseline, sex, baseline systolic and diastolic BP, and presence of diabetes.

Findings

After 7 years of follow-up, systolic BP levels among those randomised to combined HHE plus trained GP intervention were significantly lower (2.1 [4.1–0.1] mm Hg) compared to those randomised to usual care, (P = 0.04). Participants receiving the combined intervention compared to usual care had a greater reduction in LDL-cholesterol (2.7 [4.8 to 0.6] mg/dl.

Conclusions

The benefit in systolic BP reduction observed in the original cohort assigned to the combined intervention was attenuated but still evident at 7- year follow-up. These findings highlight the potential for scaling-up simple strategies for cardiovascular risk reduction in low- and middle- income countries.

Trial Registration

ClinicalTrials.gov NCT00327574     

Cloning and expression of a region of vesicle associated membrane protein2 (VAMP2) gene and its use as a recombinant peptide substrate for assaying clostridial neurotoxins in contaminated biologicals

An assay for the endopeptidase activities of clostridial neurotoxins in contaminated biotherapeutic products has been developed. Based on a synthetic peptide substrate representing amino acid residues 60–94 of the intracellular vesicle associated membrane protein2 (VAMP2), RT-PCR was used to amplify the VAMP2 sequence. The extended insert was digested with EcoRI and SalI and ligated into pGEX4T-1 vector for construction of the pGEX4T-1/VAMP plasmid for expressing in Escherichia coli a fusion protein linked to glutathione S-transferase (GST). The fusion protein was purified by affinity chromatography and used in an ELISA assay for comparison with the commercially available synthetic VAMP peptide and rabbit polyclonal antiserum. The identity of the immunoreactivity of recombinant VAMP2 protein with the chemically synthesized peptide was demonstrated by western blot. Our results indicated that recombinant VAMP2 peptide not only reacted with specific polyclonal antibody in a dose-dependent manner, without any remarkable difference observed between the reactivity of the fusion protein and commercial VAMP2 segment peptide, but also cleaved by botulinum neurotoxin type B (BONT/B) after endopeptidase assay. Thus, recombinant VAMP2 could serve as a replacement for VAMP2 synthetic peptide, potentially useful in endopeptidase assays for replacement of the currently used mouse bioassay for clostridial neurotoxins contaminating biotherapeutic products.     

Chemical diversity of essential oils from Asteriscus graveolens (Forssk.) less.: identification of cis-8-Acetoxychrysanthenyl acetate as a new natural component

Asteriscus graveolens is an endemic medicinal plant mainly distributed in south-western Algeria and south-eastern Morocco. The essential oils of leaves, stems, and flowers of A. graveolens had been studied by GC, GC/MS, and (13) C-NMR. The spectral data of two nerolidol derivatives, 6-oxo- and 6-hydroxycyclonerolidol, were reassigned by 1D- and 2D-NMR spectroscopy. These compounds can be considered as chemical markers of this genus. The structure of a monoterpenic diester with a chrysanthenane skeleton, i.e., cis-8-acetoxychrysanthenyl acetate, was determined for the first time on the basis of GC/MS, and 1D- and 2D-NMR. The stem and leaf oils were characterized by high content of oxygenated sesquiterpenes with 6-oxo- and 6-hydroxycyclonerolidol as major components, and the flower essential oils were dominated by the new monoterpenic compound cis-8-acetoxychrysanthenyl acetate.     

Synthesis and characterization of monomer and dimer complexes of porphyrin iron(III) with bridging phenylcyanamide ligands

Several new mono and dinuclear complexes of [(P)Fe^III(L)], in which P is the dianion of tetraphenylporphyrin(TPP) and tetramesitylporphyrin(TMP) and L is the monoanion of 4-azo(phenylcyanamido)benzene (apc) (1) and (2) or dianion 1,4-di cyanamidobenzene (dicyd) (3), (4), (7), (8) and 4,4′-azo-diphenylcyanamide (adpc) (5), (6), (9), (10) have been prepared by the reaction of [(P)Fe^IIICl] with appropriate thallium salts of phenylcyanamide derivatives. Each of the complexes has been characterized by FT-IR, UV-Vis, ¹H NMR, MALDI-TOF and EPR spectroscopic data. In non-coordinating solvents (such as toluene or chloroform) these complexes exhibit ¹H NMR spectra that are characteristic of high-spin (S = 5/2) species. The cyanamide group (NCN) of the bridging ligand is coordinated to Fe(III) ions through the nitrile-nitrogen. The iron(III) phenylcyanamide complexes are not reactive toward dioxygen, they convert into [TPPFe^IIICl] when treated with HCl. EPR and NMR have shown that in dinuclear complexes weak magnetic interactions take place between two iron(III) paramagnetic centers.     

Multipotent mesenchymal stem cells from adult human eye conjunctiva stromal cells

Abstract Identification of mesenchymal stem cells (MSCs) derived from alternative sources has provided an exciting prospect for intensive investigation. This work focused on characterizing a new source of MSCs from stromal cells from human eye conjunctiva. In this study, after conjunctiva biopsies and culture of stromal segment of this tissue, fibroblast-like (SH2⁺, SH3⁺, CD29⁺, CD44⁺, CD166⁺, CD13⁺) human stromal cells, which can be differentiated toward the osteogenic, adipogenic, chondrogenic, and neurogenic lineages, were obtained. These cells expressed Oct-4, Nanog, Rex-1 genes, and some lineage-specific markers like cardiac actin and Keratin. Taken together, the results indicate that conjunctiva stromal-derived cells are a new source of multipotent MSCs and despite originating from an adult source, they express undifferentiated stem cell markers.     

Process development for production of human granulocyte-colony stimulating factor by high cell density cultivation of recombinant <Emphasis Type="Italic">Escherichia coli</Emphasis>

The fed-batch process using glucose as the sole source of carbon and energy with exponential feeding rate was carried out for high cell density cultivation of recombinant Escherichia coli BL21 (DE3) expressing human granulocyte-colony stimulating factor (hG-CSF). IPTG was used to induce the expression of hG-CSF at 48 g dry cell wt l⁻¹ during high cell density culture of recombinant E. coli BL21 (DE3) [pET23a-g-csf]. The final cell density, specific yield and overall productivity of hG-CSF were obtained as ~64 g dry cell wt l⁻¹, 223 mg hG-CSF g⁻¹ dry cell wt and 775 mg hG-CSF l⁻¹ h⁻¹, respectively. The resulting purification process used cell lysis, inclusion body (IB) preparation, refolding, DEAE and Butyl-Sepharose. Effects of different process conditions such as cell lysis and washing of IB were evaluated. The results reveal that the cells lyzed at 1,200 bar, 99.9% and Triton removed about 64% of the LPS but sarcosyl had no effect on removal of nucleic acids and LPS. Further analysis show that DEAE column removes DNA about 84%. Cupper concentration was identified as parameter that could have a significant impact on aggregation, as an unacceptable pharmaceutical form that decrease process yields. The purity of purified hG-CSF was more than 99%. Also the comparison of activity between purified hG-CSF and commercial form do not show valuable decrease in activity in purified form.     

Zinc in Thalassemic Patients and Its Relation with Depression

Studies have shown that there is a relationship between zinc levels and depression. Thalassemic patients are at risk of zinc deficiency due to various causes including Desferal injection. The aim of this study, therefore, is to investigate hair zinc levels in thalassemic patients and their association with depression. For the purposes of this survey, 50 patients with major thalassemia between 10-20 years old were selected randomly. The patients' hair zinc concentration was compared with a control group of similarly aged healthy individuals. Simultaneously, their psychological status was evaluated with either the "Beck" or "Marya Kovacs" test (according to age) so that the relation between depression and zinc concentration could be assessed. The mean hair zinc concentration in patients was more than the controls (193.96 +/- 92.4 ppm vs 149.6 +/- 72.21 ppm). Zinc deficiency was present in 10% of the patients, and 52% had some degree of depression. There was a reverse correlation between zinc deficiency and blood transfusion rate (p < 0.05). Also, while there were more incidences of depression among the zinc deficient patients, the difference was not significant. Regarding the high prevalence of depression and insignificant relation to the zinc deficiency in these thalassemic patients, this research suggests the need for further consideration concerning patients' psychological status, the risk factors of zinc deficiency, as well as extended assessment into other causes of depression.     

Antioxidant defence system and physiological responses of Iranian crested wheatgrass (<Emphasis Type="Italic">Agropyron cristatum</Emphasis> L.) to drought and salinity stress

Crested wheatgrass (Agropyron cristatum L.) is a cool-season perennial grass, which has demonstrated its potential for use as turfgrass. However, limited information is available on its drought and salinity tolerance. The main purpose of this study was to investigate the changes in the antioxidant defence system and physiological traits of six Iranian crested wheatgrass genotypes under drought and salinity stresses. The experimental design comprised a split plot with water treatments (control well-watered, salinity stress and water stress) as main plots and genotypes as subplots. This study demonstrated the variations in drought and salinity tolerance among crested wheatgrass genotypes. ‘ACSKI’, ‘ACAMI’ and ‘ACDTI’ generally performed better than other genotypes under drought and salinity conditions, mainly by maintaining higher activities of antioxidant enzymes like superoxide dismutase, peroxidase, catalase, ascorbate peroxidase and non-enzyme antioxidants like glutathione, higher proline and total non-structural carbohydrates content. The increased, decreased, and unchanged activities of antioxidant enzymes in the crested wheatgrass genotypes indicates a different forms of metabolism of antioxidant enzymes in response to drought and salinity stress. In general, drought and salinity stress increases the malondialdehyde (MDA) content and hydrogen peroxide (H₂O₂) content; however, ‘ACSKI’, ‘ACAMI’ and ‘ACDTI’ genotypes could tolerate an increase in MDA content and H₂O₂ content; therefore, lower levels of MDA content and H₂O₂ content were observed. The results showed that increasing levels of diamine oxidase and polyamine oxidase have been associated with increasing drought and salinity tolerance. Based on the biochemical and physiological parameters that were evaluated, we concluded that the genotype ‘ACSKI’ was superior in terms of drought and salinity tolerance. This superiority was mainly a result of better enzymatic and non-enzymatic antioxidant defence system and better osmotic adjustment under stress conditions.