Ischemic Colitis in a Patient with Severe COVID-19 Pneumonia.

At the time of the current COVID-19 pandemic, on a daily basis, we encountered patients suffering from various manifestations of this infection. The most common are respiratory symptoms. Many of the patients require acute hospital care, and a smaller group of them are hospitalized in intensive care units. A subset of these critically ill patients demonstrates clinically remarkable hypercoagulability and thus a predisposition to venous and arterial thromboembolism, manifested by thrombotic events ranging from acute pulmonary embolism and splanchnic vascular ischemia to extremity ischemia. The article describes a case of a patient with COVID-19 pneumonia complicated by massive bleeding into the gastrointestinal tract due to ischemic enterocolitis in connection with COVID-19 infection.

Endothelial Progenitor Cells Produced From Human Pluripotent Stem Cells by a Synergistic Combination of Cytokines, Small Compounds, and Serum-Free Medium.

Human pluripotent stem cells (hPSCs) are a promising source of autologous endothelial progenitor cells (EPCs) that can be used for the treatment of vascular diseases. However, this kind of treatment requires a large amount of EPCs. Therefore, a highly efficient, robust, and easily reproducible differentiation protocol is necessary. We present a novel serum-free differentiation protocol that exploits the synergy of multiple powerful differentiation effectors. Our protocol follows the proper physiological pathway by differentiating EPCs from hPSCs in three phases that mimic in vivo embryonic vascular development. Specifically, hPSCs are differentiated into (i) primitive streak, which is subsequently turned into (ii) mesoderm, which finally differentiates into (iii) EPCs. This differentiation process yields up to 15 differentiated cells per seeded hPSC in 5 days. Endothelial progenitor cells constitute up to 97% of these derived cells. The experiments were performed on the human embryonic stem cell line H9 and six human induced pluripotent stem cell lines generated in our laboratory. Therefore, robustness was verified using many hPSC lines. Two previously established protocols were also adapted and compared to our synergistic three-phase protocol. Increased efficiency and decreased variability were observed for our differentiation protocol in comparison to the other tested protocols. Furthermore, EPCs derived from hPSCs by our protocol expressed the high-proliferative-potential EPC marker CD157 on their surface in addition to the standard EPC surface markers CD31, CD144, CD34, KDR, and CXCR4. Our protocol enables efficient fully defined production of autologous endothelial progenitors for research and clinical applications.

The Effect of Uncoated SPIONs on hiPSC-Differentiated Endothelial Cells.

BACKGROUND: Endothelial progenitor cells (EPCs) were indicated in vascular repair, angiogenesis of ischemic organs, and inhibition of formation of initial hyperplasia. Differentiation of endothelial cells (ECs) from human induced pluripotent stem cells (hiPSC)-derived endothelial cells (hiPSC-ECs) provides an unlimited supply for clinical application. Furthermore, magnetic cell labelling offers an effective way of targeting and visualization of hiPSC-ECs and is the next step towards in vivo studies. METHODS: ECs were differentiated from hiPSCs and labelled with uncoated superparamagnetic iron-oxide nanoparticles (uSPIONs). uSPION uptake was compared between hiPSC-ECs and mature ECs isolated from patients by software analysis of microscopy pictures after Prussian blue cell staining. The acute and long-term cytotoxic effects of uSPIONs were evaluated by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay) and Annexin assay. RESULTS: We showed, for the first time, uptake of uncoated SPIONs (uSPIONs) by hiPSC-ECs. In comparison with mature ECs of identical genetic background hiPSC-ECs showed lower uSPION uptake. However, all the studied endothelial cells were effectively labelled and showed magnetic properties even with low labelling concentration of uSPIONs. uSPIONs prepared by microwave plasma synthesis did not show any cytotoxicity nor impair endothelial properties. CONCLUSION: We show that hiPSC-ECs labelling with low concentration of uSPIONs is feasible and does not show any toxic effects in vitro, which is an important step towards animal studies.

Neovascularization after ischemic conditioning of the stomach and the influence of follow-up neoadjuvant chemotherapy thereon.

INTRODUCTION: Esophagectomy and reconstruction remain the optimal treatment for patients with resectable esophageal cancer. Neovascularization after ischemic conditioning of the stomach before esophagectomy is a laparoscopic procedure which may potentially reduce gastric conduit ischemia. AIM: To investigate the influence of ischemic conditioning on neovascularization along the greater curvature of the stomach and to explore the effect of neoadjuvant chemotherapy on neovascularization after ischemic conditioning. MATERIAL AND METHODS: Staging laparoscopy was performed before the main resection procedure; during this procedure ischemic conditioning was performed. Samples taken from the human stomach were divided into 3 groups: group A - patients after ischemic conditioning with a delay of 30-45 days after left gastric artery (LGA) ligation (n = 4); group B - patients who were undergoing neoadjuvant chemotherapy with a delay of 90-140 days after left gastric artery ligation (n = 4); and control group C - patients without ischemic conditioning (n = 7). RESULTS: After ischemic conditioning with a delay of 30-45 days, the count of neovessels along the greater curvature of the stomach increased from 5.4 ±0.7 in the control group to 17.5 ±0.9 in a low-power field of view (LPF) in group A and increased still further on average to 19.8 ±10.4 in group B. CONCLUSIONS: Left gastric artery ligation only is a sufficient procedure for ischemic conditioning of the stomach. Neovascularization along the greater curvature is a continuous process that depends on delay time. Neoadjuvant therapy has no influence on the effect of neovascularization.

Reprogramming of Adult Peripheral Blood Cells into Human Induced Pluripotent Stem Cells as a Safe and Accessible Source of Endothelial Cells.

New approaches in regenerative medicine and vasculogenesis have generated a demand for sufficient numbers of human endothelial cells (ECs). ECs and their progenitors reside on the interior surface of blood and lymphatic vessels or circulate in peripheral blood; however, their numbers are limited, and they are difficult to expand after isolation. Recent advances in human induced pluripotent stem cell (hiPSC) research have opened possible avenues to generate unlimited numbers of ECs from easily accessible cell sources, such as the peripheral blood. In this study, we reprogrammed peripheral blood mononuclear cells, human umbilical vein endothelial cells (HUVECs), and human saphenous vein endothelial cells (HSVECs) into hiPSCs and differentiated them into ECs. The phenotype profiles, functionality, and genome stability of all hiPSC-derived ECs were assessed and compared with HUVECs and HSVECs. hiPSC-derived ECs resembled their natural EC counterparts, as shown by the expression of the endothelial surface markers CD31 and CD144 and the results of the functional analysis. Higher expression of endothelial progenitor markers CD34 and kinase insert domain receptor (KDR) was measured in hiPSC-derived ECs. An analysis of phosphorylated histone H2AX (γH2AX) foci revealed that an increased number of DNA double-strand breaks upon reprogramming into pluripotent cells. However, differentiation into ECs restored a normal number of γH2AX foci. Our hiPSCs retained a normal karyotype, with the exception of the HSVEC-derived hiPSC line, which displayed mosaicism due to a gain of chromosome 1. Peripheral blood from adult donors is a suitable source for the unlimited production of patient-specific ECs through the hiPSC interstage. hiPSC-derived ECs are fully functional and comparable to natural ECs. The protocol is eligible for clinical applications in regenerative medicine, if the genomic stability of the pluripotent cell stage is closely monitored.

Rare cause of non-healing foot wound--acral lentiginous melanoma.

The etiology of skin integrity disorders of the lower extremities can be very diverse. In addition to common diagnoses such as venous ulcers with ischemic etiology, decubitus ulcers and diabetic foot syndrome, ulceration of a malignant etiology must also be considered. Malignant melanoma is one of the most dangerous forms of skin cancer and, although rare, may cause foot lesions. The most frequently encountered type of melanoma on acral parts of the limbs is the rare acral lentiginous variant, which only occurs in 2-10% of all melanoma types. Clinical manifestation ranges from cutaneous surface erosion to ulceration with surrounding hyperkeratosis, which can cause considerable differential diagnostic difficulties in the management of patients with non-healing foot wounds. This paper aims to present a clear case study of a Caucasian female with chronic leg ulceration that resulted in a final diagnosis of malignant acral lentiginous melanoma. Supplemental theoretical information regarding the diagnosis and treatment of malignant melanoma has also been included in the report. Malignant acral lentiginous melanoma is a rare cause of non-healing wounds, but it must be considered in cases with long-term healing complications. Precise diagnostic deliberation is crucial in the management and treatment of all chronic and long-term non-healing lesions, and appropriately performed biopsies are essential to determine whether malignancy is the primary cause.

Overexpression of c-Myb is associated with suppression of distant metastases in colorectal carcinoma.

The MYB gene codes for the c-Myb transcription factor maintaining proliferation of colon epithelial progenitors, thus controlling colon development and homeostasis. This gene is overexpressed in early phases of colorectal cancer (CRC) tumorigenesis. The aim of this study was to examine the expression of c-Myb in CRC tissue samples both at the messenger RNA (mRNA) and protein levels and to evaluate their associations with clinicopathological characteristics in a group of 108 CRC patients. Statistically significant negative association was found between the frequency of the c-Myb-positive tumor cells assessed by immunohistochemistry and the presence of distant metastases (p < 0.01) but not tumor differentiation, tumor stage, lymph node involvement, vascular invasion, tumor localization, age, and gender of the patients. Although the c-Myb protein level in the tumor tissue correlated with its mRNA level, no significant association between MYB mRNA and any clinicopathological characteristics was observed. We conclude that albeit overexpression of c-Myb is considered as an important factor contributing to early phases of CRC tumorigenesis, it may later have negative effect on tumor cell dissemination as observed recently in breast cancer as well. Further studies are required to explain the role of c-Myb during formation of CRC distant metastases.

Comparison of two non-invasive methods of microbial analysis in surgery practice: incision swabbing and the indirect imprint technique.

BACKGROUND: A variety of methods exist to take samples from surgical site infections for cultivation; however, an unambiguous and suitable method has not yet been defined. The aim of our retrospective non-randomized study was to compare two non-invasive techniques of sampling material for microbiologic analysis in surgical practice. We compared bacteria cultured from samples obtained with the use of the swab technique, defined in our study as the gold standard, with the indirect imprint technique. METHODS: A cotton-tipped swab (Copan, Brescia, Italy) was used; the imprints were taken using Whatman no. 4 filter paper (Macherey-Nagal, Duren, Germany) cut into 5×5 cm pieces placed on blood agar in a Petri dish. To culture the microorganisms in the microbiology laboratory, we used blood agar, UriSelect 4 medium (Bio-Rad, Marnes-la-Coquette, France), and a medium with sodium chloride (blood agar with salt). After careful debridement, a sample was taken from the incision surface by swab and subsequently the same area of the surface was imprinted onto filter paper. The samples were analyzed in the microbiology laboratory under standard safety precautions. The cultivation results of the two techniques were processed statistically using contingency tables and the McNemar test. Those samples that were simultaneously cultivation-positive by imprint and -negative by swabbing were processed in greater detail. RESULTS: Over the period between October 2008 and March 2013, 177 samples from 70 patients were analyzed. Sampling was carried out from 42 males and 28 females. One hundred forty-six samples were from incisions after operations (21 samples from six patients after operation on the thoracic cavity, 73 samples from 35 patients after operation on the abdominal cavity combined with the gastrointestinal tract, 52 samples from 19 patients with other surgical site infections not included above) and 31 samples from 11 patients with no post-operative infection. One patient had a sample taken both from a post-operative and a non-post-operative site. Coincidently, the most frequent cultivation finding with both techniques was a sterile one (imprint, 62; swab, 50). The microorganism cultivated most frequently after swabbing was Pseudomonas aeruginosa (22 cases), compared with Escherichia coli when the filter paper (imprint) was used (31 cases). The imprint technique was evaluated as more sensitive compared with swabbing (p=0.0001). The κ statistic used to evaluate the concordance between the two techniques was 0.302. Of the 177 samples there were 53 samples simultaneously sterile using the swab and positive in the imprint. In three samples colony- forming units (CFU) were not counted; 22 samples were within the limit of 0-25×10(1) CFU/cm(2), 20 samples within the limit of 25×10(1)-25×10(2) CFU/cm(2), five within the limit of 25×10(2)-25×10(3) CFU/cm(2), and three of more than 25×10(4) CFU/cm(2). CONCLUSIONS: The hypothesis of swabbing as a more precise technique was not confirmed. In our study the imprint technique was more sensitive than swabbing; the strength of agreement was fair. We obtained information not only on the type of the microorganism cultured, but also on the number of viable colonies, expressed in CFU/cm(2).

Study of receptor mediated selective anion transmembrane transport using parallel artificial membrane permeability assay.

The separation performance of inorganic anions ReO4(-), Br(-), SO4(2-), and NO3(-) using porphyrin-alkaloid quaternary salt transporters is examined using parallel artificial membrane permeability assay. Active and selective transport of perrhenate is achieved by porphyrin-brucine conjugates. Herein, chlorides are used as antiporters and complete removal of perrhenates from a source solution is stimulated due to electrostatic effects.

Receptor modified gold and silver nanoparticles: effect on interactions with oxoanions.

Herein we present a supramolecular non-covalent approach for the modification of gold nanoparticles (GNPs) and silver nanoparticles (SNPs) with porphyrin derivatives. The immobilization of porphyrin derivatives was carried out by two different procedures of ionic interaction. The first one was direct immobilization of the conjugate on nanoparticles and the second one was immobilization of the conjugate on 3-mercaptopropanoic acid (MPA) premodified gold nanoparticles. Such modified nanoparticles were used for interactions with selected oxoanions. The interactions were studied by UV-Vis absorption spectroscopy and electronic circular dichroism. The results showed a dependence of interaction with oxoanions on the immobilization procedures of porphyrin derivatives on the nanoparticle surface.

Modified porphyrin-brucine conjugated to gold nanoparticles and their application in photodynamic therapy.

Two porphyrin-brucine quaternary ammonium salts were immobilized on gold nanoparticles and their suitability for both in vitro and in vivo photodynamic therapy (PDT) was assayed using the basaloid squamous cell carcinoma PE/CA-PJ34 cell line. In vitro PDT experiments revealed that the gold nanoparticle-bound conjugates were less effective than unbound conjugates in killing cells. However, the same conjugates were more effective in reducing tumor size in vivo, with complete tumor regression observed.