The Angiopoietin Signaling Pathway Is Involved in Inflammatory Processes in Hospitalized COVID-19 Patients.

The viral agent SARS-CoV-2 clearly affects several organ systems, including the cardiovascular system. Angiopoietins are involved in vascular integrity and angiogenesis. Angiopoietin-1 (Ang1) promotes vessel stabilization, while angiopoietin-2 (Ang2), which is usually expressed at low levels, is significantly elevated in inflammatory and angiogenic conditions. Interleukin-6 (IL-6) is known to induce defective angiogenesis via the activation of the Ang2 pathway. Vasculitis and vasculopathy are some of the defining features of moderate to severe COVID-19-associated systemic disease. We investigated the serum levels of angiopoietins, as well as interleukin-6 levels and anti-SARS-CoV2 IgG titers, in hospitalized COVID-19 patients across disease severity and healthy controls. Ang2 levels were elevated in COVID-19 patients across all severity compared to healthy controls, while Ang1 levels were decreased. The patients with adverse outcomes (death and/or prolonged hospitalization) had relatively lower and stable Ang1 levels but continuously elevated Ang2 levels, while those who had no adverse outcomes had increasing levels of both Ang1 and Ang2, followed by a decrease in both. These results suggest that the dynamic levels of Ang1 and Ang2 during the clinical course may predict adverse outcomes in COVID-19 patients. Ang1 seems to play an important role in controlling Ang2-related inflammatory mechanisms in COVID-19 patients. IL-6 and anti-SARS-CoV2 spike protein IgG levels were significantly elevated in patients with severe disease. Our findings represent an informative pilot assessment into the role of the angiopoietin signaling pathway in the inflammatory response in COVID-19.

Ephrin-A1 and the sheddase ADAM12 are upregulated in COVID-19.

More than 3.5 million people have died globally from COVID-19, yet an effective therapy is not available. It is, therefore, important to understand the signaling pathways that mediate disease progression in order to identify new molecular targets for therapeutic development. Here, we report that the blood serum levels of ephrin-A1 and the sheddase ADAM12 were significantly elevated in COVID-19 patients treated at SUNY Downstate Hospital of Brooklyn, New York. Both ephrin-A1 and ADAM12 are known to be involved in inflammation and regulate endothelial cell permeability, thus providing a gateway to lung injury. The clinical outcome correlated with the ephrin-A1 and ADAM12 serum levels during the first week of hospitalization. In contrast, the serum levels of TNFα were elevated in only a small subset of the patients, and these same patients also had highly elevated levels of the sheddase ADAM17. These data indicate that ephrin-A1-mediated inflammatory signaling may contribute to COVID-19 disease progression more so than TNFα-mediated inflammatory signaling. They also support the notion that, in COVID-19 inflammation, ADAM12 sheds ephrin-A1, while ADAM17 sheds TNFα. Furthermore, the results suggest that elevated serum levels and activity of cytokines, such as TNFα, and other secreted inflammatory molecules, such as ephrin-A1, are not simply due to overexpression, but also to upregulation of sheddases that release them into the blood circulation. Our results identify ephrin-A1, ADAM12, and other molecules in the ephrin-A1 signaling pathway as potential pharmacological targets for treating COVID-19 inflammation.

Ultrarapid EGFR Mutation Screening Followed by Comprehensive Next-Generation Sequencing: A Feasible, Informative Approach for Lung Carcinoma Cytology Specimens With a High Success Rate.

INTRODUCTION: For patients with advanced NSCLC, cytologic samples may be the only diagnostic specimen available for molecular profiling. Although both rapid and comprehensive assessment are essential in this setting, an integrated multitest approach remains an important strategy in many laboratories, despite the risks and challenges when working with scant samples. In this study, we describe our experience and high success rate in using a multitest approach, focusing on the clinical validation and incorporation of ultrarapid EGFR testing using the Idylla system followed by comprehensive next-generation sequencing (NGS). METHODS: Cytology samples received for routine molecular testing were included in this study. The performance characteristics of the EGFR Idylla assay were assessed; tissue suitability parameters and interpretation criteria to supplement automated mutation calling were established. The assay performance was monitored for 1 year, comparing the results with those of concurrent NGS testing by MSK-IMPACT (primarily) or MSK-AmpliSeq and MSK-Fusion solid panel in a subset of cases. RESULTS: Overall, 301 samples were studied; 83 samples were included in validation (60.2% [50 of 83] were positive for EGFR mutations). Concordance with the reference method was 96.4% (80 of 83) of the samples with excellent reproducibility. The limit of detection was variable depending on the total tissue input and the specific mutation tested. Unextracted tissue inputs that maintained total EGFR cycle of quantification at less than 23 allowed all mutations to be detected if present at greater than 5% variant allele frequency. Mutations could be detected at 1% variant allele frequency with total EGFR cycle of quantification of 18. During the clinical implementation phase, 218 NSCLC samples were tested by Idylla (24.3% [53 of 218] were EGFR mutation positive). Concurrent NGS testing was requested on 165 samples and successfully performed on 96.4% (159 of 165) of the samples. The Idylla automated results were concordant with those obtained by NGS in 96.2% (153 of 159) of cases and improved to 98.7% (157 of 159) after incorporation of manual review criteria to supplement automated calling, resulting in a diagnostic sensitivity of 95.6% (95% confidence interval: 84.9%-99.5%). In general, 9% (14 of 159) of the cases tested by NGS had EGFR mutations not covered by the Idylla assay, primarily insertions in exon 19 and 20 and minor mutations cooccurring with canonical sensitizing mutations. CONCLUSIONS: Comprehensive molecular testing is feasible and has a high success rate in NSCLC cytology samples when using a multitest approach. Testing with the Idylla system enables rapid and accurate determination of the EGFR status without compromising subsequent NGS testing.

A Comprehensive, Epidemiological and Ecological Descriptive Study on Vitamin D Status in Iran (308005 People, from 2009-2018).

BACKGROUND: Vitamin D is an essential substance for absorption of calcium and phosphorus from intestine so it is vital for muscles and skeletal development. Deficiency of this vitamin is pandemic. The vitamin D status depends on the different factors such as UV exposure, diet, and ecological features of living location, age and gender. The aim of this study was to describe the vitamin D level in different provinces of Iran and to investigate the association between vitamin D status and multiple variables. METHODS: We collected the serum 25(OH)D (Vitamin D) level data of 308,005 people referred to different laboratories from 30 provinces of Iran and organized them by each province, year, age, gender, precipitation, latitude and longitude, and humidity over 10 yr (2009-2018). Data were analyzed to find out the correlation between age, gender, longitude and latitude, humidity and sum of precipitation. RESULTS: West Azerbaijan had the highest level of vitamin D with a mean level of 33.24 and a standard deviation of 32.001, and North Khorasan had the lowest level with a mean level of 14.46 and a standard deviation of 8.980 among 30 provinces of Iran. The correlation between all studied variables (age, and gender, latitude and longitude, humidity, the sum of precipitation) was significant (P<0.001). CONCLUSION: The average total vitamin D level in Iran is 25.41 ng/ml, which is within the area of deficiency. Vitamin D is associated with age, and gender, latitude and longitude, humidity, the sum of precipitation. So changes in any of these variables can lead to vitamin D alteration.

Major Discrepancies of HCV Viral Loads Determined by Real Time Polymerase Chain Reaction Caused by Use of Collection Tubes that Differ Only with the Presence or Absence of Serum Separator.

BACKGROUND: For viral load (VL) analysis of hepatitis C virus (HCV) by Real Time PCR, approved collection tubes were lavender top (LTT) and plasma processing (PPT) tubes, that differ in that PPTs include plasma separator. Using a Real Time PCR method, we investigated how the results correlated for the two tube types. METHODS: The plasma fractions of blood samples from each of 202 patients were collected in LTT and PPT tubes at the same time and were assayed for HCV VL by the Abbott m2000 Real Time PCR System; the results were analyzed statistically. RESULTS: VLs for 103 samples for both tubes were negative. Positive results were obtained for another 79 tube pairs, including six with VLs for LTTs but values below linearity for the corresponding PPTs. For the 73 samples for which quantitative results were obtained for both tube types, VLs were statistically higher in LTTs (means 1,817,821.8 in LTTs and 1,083,669.1 in PPTs, p=0.006, alpha=0.05) using the paired t-test and confirmed by the Chi Square and McNemars tests. CONCLUSIONS: VLs in LTTs are significantly higher and more sensitive than in PPTs, suggesting the necessity of standardization of collection tubes for HCV VLs.

Do Serum Creatinine Levels Show Clinically Significant Fluctuations on Serial Determinations on the Siemens Advia 1800 Analyzer?

BACKGROUND: The goal of this work was to determine whether there are clinically significant fluctuations in the level of serum creatinine on serial determinations, especially in the borderline range (1.1-1.3 mg/dl), after specimen storage. METHODS: Sixty-one serum samples were analyzed. They were divided into three categories based on the initial serum creatinine measurement: low (≤1.0 mg/dl), borderline (1.1-1.3 mg/dl), and high (≥1.4 mg/dl). The specimens were stored at 4°C and run on the Siemens Advia 1800 chemistry analyzer on days 1, 3, and 11. RESULTS: Statistical comparisons of the three groups were made using the unpaired t-test, yielding a two-tailed P-value for each group comparison. The P-values ranged from 0.0829 to 0.3892, indicating no statistically significant difference between the standard deviations of each group. CONCLUSIONS: Mild-to-moderate fluctuations in precision occur in successive serum creatinine determinations. The overwhelming majority of these fluctuations should not affect clinical decision making.

Designing a prognostic scoring system for predicting the outcomes of proximal fifth metatarsal fractures at 20 weeks.

BACKGROUND: Fractures of the proximal fifth metatarsal bone are among the most common fractures observed in the foot and their classification and management has been subject to much discussion and disagreement. In this study, we aim to identify and quantify the effect of possible predictors of the outcome of the treatment of proximal fifth metatarsal fractures. METHODS: Patients with established proximal fifth metatarsal fractures were enrolled in this prospective cohort and the outcome of their treatment was assessed using the AOFAS mid foot scale at 6 and 20 weeks. RESULTS: 143 patients were included in the study. Our study showed that displacement, weight and type III fractures were significant independent predictors of poor outcome at 6 weeks while at 20 weeks in addition to these factors, gender and diabetes mellitus were also shown to be significant independent predictors of poor outcome. A scoring system was designed by assigning weight to these factors and it was shown to be a strong predictor of outcome at 20 weeks. CONCLUSION: We recommend that our scoring system would help surgeons to decide whether patients' prognostic factors are significant enough for him/her to opt for a surgical approach to treatment rather than a conservative approach.

Overexpression of signal transducers and activators of transcription in embryos derived from vitrified oocytes negatively affect E-cadherin expression and embryo development.

Vitrification apart from all drawbacks on oocyte ultra-structure can affect the oocyte mRNA content. Among those evaluated transcripts, no data is available regarding the effect of vitrification on signal transducer and activator of transcription (STAT3) expression in oocytes and the resulting preimplantation embryos. Considering the bidirectional relationship between E-cadherin (CDH1) and STAT3 and the adverse effect of cryopreservation on adherent junctions, we aimed to ascertain to what extent STAT3 and CDH1 genes expression is affected by vitrification in oocytes and the resulting embryos. The ovine vitrified-warmed and fresh GV oocytes were separately subjected to in vitro maturation and fertilization and cultured up to the blastocyst stage. The relative abundance of STAT3 and CDH1 transcripts were analysed by RT-PCR in both classes of fresh and vitrified GV and MII oocytes and the resulting embryos at 2-7 cells, 8-16 cells, morula, and blastocyst stages. Vitrified oocytes showed lower cleavage (37.8% vs. 95.9%, P<0.001) and blastocyst (8.1% vs. 52.7%, P<0.001) rates compared to control. The relative mRNA abundance of both genes was increased after oocyte maturation indicating their expression was started earlier than expected time proposed for embryonic genomic activation. In embryos derived from both fresh and vitrified oocytes, the maximum concentrations of STAT3 and CDH1 transcripts were observed at 2-7 cells and morula stages, respectively. Moreover, in contrast to CDH1 the relative expression of STAT3 in vitrified derived embryos was higher than embryos derived from fresh oocytes. The overexpression of STAT3 in embryos derived from vitrified oocytes might be the reason for the lower CDH1 expression and in turn the lower developmental competence of the resulting embryos.

A randomized clinical trial on comparison of corticosteroid injection with or without splinting versus saline injection with or without splinting in patients with lateral epicondylitis.

BACKGROUND: Lateral epicondylitis is a common problem affecting 1-3% of the population. There has been much debate about the best treatment modality for this condition. There is, however, no conclusive evidence in support of any of the proposed treatment modalities. In this trial, we have studied the effect of corticosteroid injection (with or without splinting) with normal saline injection (with or without splinting). MATERIALS AND METHODS: In this double-blind, randomized clinical trial, individuals were randomly assigned to either of four treatment groups and received either 40 mg depomedrol injection alone, 40 mg depomedrol injection with splinting, normal saline injection alone, or normal saline injection with splinting. They were evaluated using the visual analog scale (VAS) at weeks 2, 4 and 24 and with the Oxford elbow scale (OES) at 24 weeks. RESULTS: A total of 79 patients were participated in the study. The corticosteroid injection groups had better pain relief as measured by VAS at 2 and 4 weeks compared with the two saline injection groups. Mean VAS difference at week 0 versus week 2 was 4.5 ± 0.9 and 2.8 ± 0.6 in corticosteroid injection groups and saline injection groups respectively (P < 0.01) but at 24 weeks, there was only moderate benefit reported for the group which received steroid injection and splinting (P < 0.01) compared to the saline injection groups. The saline injection groups reported better improvement in OES scores (20.1 ± 3.7) at the end of the trial compared corticosteroid injection groups (16.1 ± 2.9) (P < 0.05). CONCLUSION: Our results indicate that despite the clear pain reduction benefit associated with steroid injection in short term, this benefit in comparison with normal saline injection fades by the 24(th) week of follow-up.

Development of liposomes loaded with anti-leishmanial drugs for the treatment of cutaneous leishmaniasis.

Cutaneous leishmaniasis is caused by different species of Leishmania parasites and its available treatments have not yet provided a strong consistent result. The weak response of current chemotherapeutics is due to their deficient effects on stealth parasites inside macrophages, rapid clearance from the site of action and systemic side effects in high doses. Liposomal formulation of anti-leishmanial drugs could overcome these problems. In this study, different liposomal formulations of three famous anti-leishmanial drugs: Glucantime®, miltefosine and paromomycin were prepared by a modified freeze-drying double emulsion method. Liposome size, zeta potential and encapsulation efficiency were evaluated, and their imaging was carried out by means of atomic force microscopy. Three formulations were evaluated in vivo by subcutaneous injection into skin lesions caused by Leishmania major in BALB/c mice. Encapsulation efficiency of prepared liposomes was up to 90%; however, they inherited a bimodal size distribution that caused their encapsulation efficiency to decrease to 50% during filtering sterilization. Besides, the effect of surface charge was significant on preparation procedure, size and encapsulation efficiency. All three formulations reduced amastigote counts and lesion size but only miltefosine-loaded formulations had significant therapeutic effects compared with control group (p < 0.05).

Results of a unilateral lip lift for correction of a vertical disproportion in upper lip vascular anomalies.

Congenital vascular anomalies most notably hemangiomas involving the lips, especially those which fail to regress, pose a difficult problem both for the surgeon and the patient. These lesions not only discolour the skin but may also distort the shape of the lip. When nonsurgical modalities fail to treat these lesions, surgery is chosen as the next viable step. In this article, we propose a well planned sequential surgical procedure for unilateral lesions. This approach in comparison with previously used procedures produces noticeably better results and fewer complications. During a period of 4 years, we treated 21 patients with vascular anomalies using the unilateral upper lip lift procedure to correct the vertical disproportion of commissures. Using this procedure, we managed to restore the normal form and symmetry of the upper lip in a majority of our patients with less obvious scarring and few complications.

Adenoviral gene therapy in gastric cancer: a review.

Gastric cancer is one of the most common malignancies worldwide. With current therapeutic approaches the prognosis of gastric cancer is very poor, as gastric cancer accounts for the second most common cause of death in cancer related deaths. Gastric cancer like almost all other cancers has a molecular genetic basis which relies on disruption in normal cellular regulatory mechanisms regarding cell growth, apoptosis and cell division. Thus novel therapeutic approaches such as gene therapy promise to become the alternative choice of treatment in gastric cancer. In gene therapy, suicide genes, tumor suppressor genes and anti-angiogenesis genes among many others are introduced to cancer cells via vectors. Some of the vectors widely used in gene therapy are Adenoviral vectors. This review provides an update of the new developments in adenoviral cancer gene therapy including strategies for inducing apoptosis, inhibiting metastasis and targeting the cancer cells.

The use of oral cyclophosphamide with dexamethasone pulse therapy in the treatment of pemphigus vulgaris.

BACKGROUND: Pemphigus vulgaris (PV) is a severe acquired autoimmune disease. During the past 15 years we used dexamethasone pulse therapy (DP) and oral cyclophosphamide for treatment of our patients with PV. OBJECTIVES: To evaluate the efficacy and side effects of this treatment. METHOD: The study was a retrospective study. Medical records of 50 known cases of PV, who were treated with dexamethasone pulse therapy and oral cyclophosphamide were evaluated and the patients were re-examined and questioned about their current situations. RESULTS: Thirty-five females and 15 males were treated, out of which 38 finished the study. At the end of study, 21 patients were in remission, 10 patients were in the healing stage, five patients were partially healed and two patients died. The side effects of this method, especially the adverse effects of cyclophosphamide, were less than previously reported. CONCLUSION: Dexamethasone pulse therapy and oral cyclophosphamide is an effective regimen with few side effects in the treatment of mild to moderate PV.