Red cell and platelet‐derived microparticles are increased in G6PD‐deficient subjects

In response to oxidative stress and during apoptosis, cells often shed microparticles (MPs), submicron elements carrying phosphatidylserine and protein antigens. Glucose‐6‐phosphate dehydrogenase (G6PD)‐deficient cells are extremely sensitive to oxidative damage that may lead to the formation of MPs. To determine whether G6PD deficiency alters membrane phospholipid asymmetry and increases MPs production, we determined the concentrations and cellular origins of MPs in G6PD‐deficient individuals using flow cytometry. G6PD‐deficient individuals showed an increase in circulating MPs concentrations as compared with G6PD‐normal individuals [1051/μL (865–2532/μL) vs. 258/μL (235–575/μL), P < 0.01]. MPs concentrations were significantly increased with the severity of G6PD deficiency. Median MPs concentrations from individuals with severe G6PD deficiency, and individuals with moderate G6PD deficiency were 2567/μL (1216–2532/μL) and 984/μL (685–2107/μL), respectively (P < 0.01). Importantly, G6PD enzymatic activity was significantly correlated with MPs concentrations with r² = 0.731. MPs found in G6PD deficiency individuals were largely derived from red blood cells (RBCs) (45%) and platelets (30%). Additionally, Atomic Force Microscopy was used to study the morphology and measures the diameter of MPs found in G6PD‐deficient individuals. The mean (SD) width and height of RMPs were 0. 41 (0.18) and 2.04 (0.14) μm, respectively. Together, these results indicate that MP concentration is significantly correlated with G6PD enzymatic activity and is increased in G6PD‐deficient as compared with G6PD‐normal individuals. Our data also provide an evidence for an alteration in cell membrane associated with a decreased in G6PD activity. However, the significance of MPs in G6PD deficiency needs further clarification.     

Polymeric nano- and microparticle technologies for oral gene delivery

Gene therapy refers to local or systemic administration of a nucleic acid construct that can prevent, treat and even cure diseases by changing the expression of genes that are responsible for the pathological condition. Oral gene therapy has significant promise for treatment of local diseases such as inflammatory bowel disease and for systemic absorption of the expressed protein therapeutics. In addition, efficient oral delivery of DNA vaccines can have significant impact in disease prevention. The use of polymeric gene delivery vectors promises the translation of this experimental medical concept into clinical reality. This review addresses the challenges and opportunities in the development of polymer-based nano- and microparticle technologies for oral gene therapy. Specifically, the discussion is focused on different synthetic and natural polymers used for formulating nano- and microparticle technologies and the use of these delivery systems for oral DNA administration for therapeutic and vaccination purposes.     

Significance of increasing adhesion of cord blood hematopoietic cells and a new method: platelet microparticles

Hematopoietic recovery after transplantation with cord blood (CB) is slower than with bone marrow (BM) and mobilized peripheral blood. Adhesion molecules (AMs) on hematopoietic cells are involved in hematopoietic cells' homing. It may be possible to enhance CB CD34+ cells engraftment by increasing their expressions of AM. Twenty-three patients with childhood acute leukemia treated with unrelated CBT were studied. It was found that the time to neutrophil recovery correlated with CXCR4 and the time to platelet recovery correlated with both CD62L and CXCR4. Platelet microparticles (PMPs) carry some AMs such as aIIb b (CD41), P-selectin (CD62P), and CXCR4, CD34+ cells express platelet-binding antigens (CD162 and CD11b). It was found that AMs were increased dramatically on CD34+ cells surface in the presence of PMPs, and CD34+ cells covered with PMPs adhered better to human umbilical vein endothelial cells and fibronectin. These findings suggested that PMPs could increase adhesion of donor's cells to host BM in CBT.     

Toxicological implications of amplifying the antibacterial activity of gallic acid by immobilisation on silica particles: A study on C. elegans

Immobilisation of natural compounds on solid supports to amplify antimicrobial properties has reported successful results, but modifications to physico-chemical properties can also imply modifications from a toxicological viewpoint. This work aimed to study the immobilising process of gallic acid in the antibacterial activity of L. innocua and its toxicological properties in vivo using Caenorhabditis elegans. The experiment was based on obtaining the minimum bactericidal concentration for free and immobilised gallic acid by comparing lethality, locomotion behaviour, chemotaxis and thermal stress resistance on C.elegans at those concentrations. The results showed a lowering minimum bactericidal concentration and modifications to nematode responses. Increased lethality and velocity of movements was observed. Immobilisation increased the repellent effect of gallic acid with a negative chemotaxis index. Thermal stress resistance was also affected, with higher mortality for immobilised gallic acid compared to bare particles and free gallic acid. Thus despite evidencing a generalised increase in the toxicity of gallic acid in vivo, lowering the minimum bactericidal concentration allowed a bacterial reduction of 99 % with less than one third of mortality for the nematodes exposed to free gallic acid.     

The Evaluation of Leukocytes in Response to the In Vitro Testing of Ventricular Assist Devices

Infection is a clinically relevant adverse event in patients with ventricular assist device (VAD) support. The risk of infection could be linked to a reduced immune response resulting from damage to leukocytes during VAD support. The purpose of this study was to develop an understanding of leukocyte responses during the in vitro testing of VADs by analyzing the changes to their morphology and biochemistry. The VentrAssist implantable rotary blood pump (IRBP) and RotaFlow centrifugal pump (CP) were tested in vitro under constant hemodynamic conditions. Automated hematology analysis of samples collected regularly over 25‐h tests was undertaken. A new flow cytometric assay was employed to measure biochemical alteration, necrosis (7‐AAD) and morphological alteration (CD45 expression) of the circulating leukocytes during the pumping process. The results of hematology analysis show the total leukocyte number and subset counts decreased over the period of in vitro tests dependent on different blood pumps. The percentage of leukocytes damaged during 6‐h tests was 40.8 ± 5.7% for the VentrAssist IRBP, 17.6 ± 5.4% for the RotaFlow CP, and 2.7 ± 1.8% for the static control (all n = 5). Flow cytometric monitoring of CD45 expression and forward/side scatter characteristics revealed leukocytes that were fragmented into smaller pieces (microparticles). Scanning electron microscopy and imaging flow cytometry were used to confirm this. Device developers could use these robust cellular assays to gain a better understanding of leukocyte‐specific VAD performance.     

HLA Antigens Shed from the Surface of Synthetic or Naturally Occurred Platelet-Derived Microparticles During Storage of Platelet Concentrate

The demand for standard platelet concentrates (PCs) has continued to increase in the recent years. Infusible platelet membranes (IPM) prepared from new or outdated human platelets have been developed as an alternative to standard PCs, with the additional advantage of long shelf life and increased viral safety. Reduction of HLA antigens on the IPM has been assigned as one of the probable advantages of this product. In re-examining this issue, we studied the existence of HLA class I on the surface of IPM microparticles. In comparison we also surveyed HLA expression on the surface of the naturally occurred platelet-derived microparticles (nPMPs) during 7 days storage. Intended for producing IPM, PCs obtained from Iranian blood transfusion organization were lysed; virally inactivated with wet heat in the presence of a heat stabilizer and then sonicated. IPMs were separated using centrifugation and liquid-stored in 4°C. The expression of HLA class I antigens was surveyed using flow cytometry technique. HLA molecules were present on the microparticles. Shedding of HLA antigens was demonstrated from the surface of the both liquid-stored IPM and nPMPs during storage. Storage of IPM in 4°C was accompanied with significant reduction of HLA molecules. It seemed that achievement of HLA-free IPM could be impossible unless chloroquine treated platelets were used to prepare these microvesicles.     

Doxycycline hydrochloride-metronidazole solid lipid microparticles gels for treatment of periodontitis: development,in-vitro and in-vivo clinical evaluation

Objective: To formulate solid lipid microparticles (SLMs) encapsulating doxycycline hydrochloride (DH) and metronidazole (MT) for the treatment of periodontal diseases.

Methods: SLMs were prepared applying hot homogenization method, using different types of lipids and stabilized with various types and concentrations of surfactants. The optimized formula was subjected to freeze-drying followed by incorporation into poloxamer gel. Microbiological and clinical evaluation of the selected SLMs on patients suffering from periodontal diseases was performed.

Results: SLMs could entrap high percentage of both drugs (81.14% and 68.75 % for doxycycline hydrochloride and metronidazole respectively). Transmission electron microscopy images of SLMs showed nearly spherical particles. Freeze-dried SLMs showed satisfactory stability for three months. Combined drugs were molecularly dispersed in SLMs. Incorporation of the freeze-dried SLMs powder in poloxamer gel could control the drugs release for 72 h. In-vivo study revealed effective and safe use of SLMs gel for periodontitis treatment. Significant improvement in both microbiological and clinical parameters was observed as compared to scaling and root planing alone.

Conclusion: The formulated SLMs gel offers an applicable dosage form that can be injected directly into the periodontal pocket as adjunctive to scaling and root planing.     

Evaluation of Glass Delamination Risk in Pharmaceutical 10 mL/10R Vials

Glass delamination is characterized by the dissociation of glass flakes from the glass surface. Since glass delamination is time dependent, 5 vial types were investigated to assess delamination under accelerated stress conditions published as quick tests in literature and compared to stress testing recommended per United States Pharmacopoeia <1660>. A broad panel of analytical techniques was employed to test the solution for visible/subvisible particles and leachables and characterize topography and composition of the surface. The vial types showed significant differences in surface durability when applying the same stress conditions. An increase in glass leachables and change in topography were shown for uncoated vials. An indication for an elevated delamination risk was confirmed for Expansion 33 vials only by the compiled analytical data set including particle assessment and change in elemental composition of the near glass surface investigated by dynamic secondary ion mass spectrometry. The delamination test protocols differ in test solution, handling, and time. Before choosing the most appropriate protocol to predict delamination propensity and mimic real-time conditions, long-term storage data are needed. A combination of analytical techniques to study the risk for long-term corrosion of glass is highly recommended covering the 3 aspects: visible/subvisible particle assessment, solution analysis, and surface characterization.