Parker Solar Probe Observations of Solar Wind Energetic Proton Beams Produced by Magnetic Reconnection in the Near-Sun Heliospheric Current Sheet.

We report observations of reconnection exhausts in the Heliospheric Current Sheet (HCS) during Parker Solar Probe Encounters 08 and 07, at 16 R s and 20 R s , respectively. Heliospheric current sheet (HCS) reconnection accelerated protons to almost twice the solar wind speed and increased the proton core energy by a factor of ∼3, due to the Alfvén speed being comparable to the solar wind flow speed at these near-Sun distances. Furthermore, protons were energized to super-thermal energies. During E08, energized protons were found to have leaked out of the exhaust along separatrix field lines, appearing as field-aligned energetic proton beams in a broad region outside the HCS. Concurrent dropouts of strahl electrons, indicating disconnection from the Sun, provide further evidence for the HCS being the source of the beams. Around the HCS in E07, there were also proton beams but without electron strahl dropouts, indicating that their origin was not the local HCS reconnection exhaust.

Association of intraoperative hypotension with postoperative morbidity and mortality: systematic review and meta-analysis.

BACKGROUND: Intraoperative hypotension, with varying definitions in literature, may be associated with postoperative complications. The aim of this meta-analysis was to assess the association of intraoperative hypotension with postoperative morbidity and mortality. METHODS: MEDLINE, Embase and Cochrane databases were searched for studies published between January 1990 and August 2018. The primary endpoints were postoperative overall morbidity and mortality. Secondary endpoints were postoperative cardiac outcomes, acute kidney injury, stroke, delirium, surgical outcomes and combined outcomes. Subgroup analyses, sensitivity analyses and a meta-regression were performed to test the robustness of the results and to explore heterogeneity. RESULTS: The search identified 2931 studies, of which 29 were included in the meta-analysis, consisting of 130 862 patients. Intraoperative hypotension was associated with an increased risk of morbidity (odds ratio (OR) 2.08, 95 per cent confidence interval 1.56 to 2.77) and mortality (OR 1.94, 1.32 to 2.84). In the secondary analyses, intraoperative hypotension was associated with cardiac complications (OR 2.44, 1.52 to 3.93) and acute kidney injury (OR 2.69, 1.31 to 5.55). Overall heterogeneity was high, with an I2 value of 88 per cent. When hypotension severity, outcome severity and study population variables were added to the meta-regression, heterogeneity was reduced to 50 per cent. CONCLUSION: Intraoperative hypotension during non-cardiac surgery is associated with postoperative cardiac and renal morbidity, and mortality. A universally accepted standard definition of hypotension would facilitate further research into this topic.

DNA methylation profiles of immune response-related genes in apical periodontitis.

AIM: To investigate the DNA methylation profiles of immune response-related genes in apical periodontitis (AP) lesions. METHODOLOGY: The methylation profiles on the cytosine-phosphate-guanine (CpG) regions of 22 gene promoters involved in inflammation and autoimmunity were assessed in 60 human AP lesions and 24 healthy periodontal ligaments (controls) using a pathway-specific real-time polymerase chain reaction array (EpiTect® Methyl Signature PCR Array Human Inflammatory Response). Differentially methylated genes were subsequently assessed for their mRNA expression. Data analyses (One-way anova, Tukey's multiple comparisons tests and Mann-Whitney tests) were performed using GraphPad Prism 6 software. P values ≤ 0.05 were considered statistically significant. RESULTS: Significant DNA hypermethylation was observed for CXCL3 and FADD gene promoters in AP lesions when compared to control tissues (P < 0.001) and among other genes (P < 0.05). In contrast, IL12B and IL4R were associated with significant hypomethylation in comparison to other genes (P < 0.05). IL12B, IL4R, CXCL3 and FADD had differential mRNA expression in AP lesions and controls (P < 0.001). CONCLUSIONS: Differential methylation profiles of immune response-related genes, such as FADD, CXCL3, IL12B and IL4R, may have an influence on individual AP susceptibility and patient treatment outcomes, through their potential contributions to altered expression of disease-relevant genes. Methylation and/or genetic variations in additional genes may also contribute to the dynamics of AP development and should be considered in future studies.

The association of hypotension during non-cardiac surgery, before and after skin incision, with postoperative acute kidney injury: a retrospective cohort analysis.

Intra-operative hypotension is associated with acute postoperative kidney injury. It is unclear how much hypotension occurs before skin incision compared with after, or whether hypotension in these two periods is similarly associated with postoperative kidney injury. We analysed the association of mean arterial pressure < 65 mmHg with postoperative kidney injury in 42,825 patients who were anaesthetised for elective non-cardiac surgery. Intra-operative hypotension occurred in 30,423 (71%) patients: 22,569 (53%) patients before skin incision; and 24,102 (56%) patients after incision. Anaesthetised patients who were hypotensive had mean arterial pressures < 65 mmHg for a median (IQR [range]) of 5.5 (0.0-14.7 [0.0-60.0]) min.h-1 before skin incision, compared with 1.7 [0.3-5.1 [0.0-57.5]) min.h-1 after incision: a median (IQR [range]) of 36% (0%-84% [0%-100%]) of hypotensive readings were before incision. We diagnosed postoperative kidney injury in 2328 (5%) patients. The odds ratio (95%CI) for acute kidney injury was 1.05 (1.02-1.07) for each doubling of the duration of hypotension, p < 0.001. Postoperative kidney injury was associated with the product of hypotension duration and severity, that is, area under the curve, before skin incision and after, odds ratio (95%CI): 1.02 (1.01-1.04), p = 0.004; and 1.02 (1.00-1.04), p = 0.016, respectively. A substantial fraction of all hypotension happened before surgical incision and was thus completely due to anaesthetic management. We recommend that anaesthetists should avoid mean arterial pressure < 65 mmHg during surgery, especially after induction, assuming that its association with postoperative kidney injury is, at least in part, causal.

Monitoring whole-lung lavage using lung ultrasound: The changing phases of the lung.

Lung ultrasound (LUS) has been proven to yield valuable information for lung and pleural pathology. It is well validated for assessing extravascular lung water. It can also be used to monitor stages of controlled lung de-aeration in whole lung lavage (WLL) which is the treatment for Pulmonary Alveolar Protienosis (PAP),characterized by abnormal surfactant in the alveoli affecting gas exchange .LUS can help decide the point of termination of lung flooding. A 55 year old lady with biopsy proven pulmonary alveolar proteinosis presented with respiratory failure. WLL was planned. LUS was used to study the stages of lung flooding as previously described for ARDS model.6 areas screened based on six areas that are normally examined like upper zone, mid zone and lower zone showed alveolar interstitial pattern. One lung ventilation (OLV) was done and isolation of lavage lung was confirmed which was seen as lung collapse (lung pulse) on LUS. Saline infusion resulted in increase in B lines followed by tissue like pattern with fluid bronchogram on LUS(alveolar flooding) in all the areas. During the lavage of the second lung, appearance of alveolar flooding pattern resulted in termination of saline infusion. The use of LUS in monitoring WLL reduced amount of saline used for lavage, pick up complications like pleural effusion and spillage.

Efficient Purification of rhG-CSF and its PEGylated Forms and Evaluation for In Vitro Activities.

Granulocyte-colony stimulating factor (G-CSF) has commonly been used to help the patients to recover from neutropenia inflicted due to radiotherapy, organ transplants and chemotherapy. As the number of people undergoing these therapies and procedures are increasing world-wide, the need for more economical ways of G-CSF production and improvement in its efficacy has become increasingly crucial. In the present study, recombinant human G-CSF (rhG-CSF) was expressed in E. coli and its purification process was optimized by demonstrating better efficiency and higher recoveries (upto 54%) in a multi-step chromatographic purification process, which is greater than the existing reports. Additionally, the efficacy of rhG-CSF was increased by derivatizing with polyethylene glycol (PEG; upto 85% PEGylation), which increases the plasma clearance time, reduces the immunogenicity and requires less frequent administration to the patient. Overall, the present study suggests a cost-effective purification process of rhG-CSF and also proposes its efficient conjugation with PEG for enhanced efficacy as compared to the existing commercially available forms.

Cystatin C: An alternative dialysis adequacy marker in high flux hemodialysis.

The conventional, low flux (LF) dialyzer allows the removal of small molecular solutes like urea and creatinine. High flux (HF) dialyzers allow the effective removal of middle molecules (MM) as well, and are associated with reduced hemodialysis-related morbidity and mortality. Cystatin C has attractive characteristics as a representative MM. The aim of this study was to determine cystatin C reduction ratio (CysCRR) in both LF and HF groups and to compare it with other markers of dialysis adequacy. Thirty-seven patients were subjected to both LF and HF hemodialysis 2 weeks apart. Serum urea, creatinine and cystatin C were measured pre- and post-dialysis. Cystatin C was measured by latex-enhanced immunoturbidimetry. Urea and creatinine reduction ratios were 72.3 ± 14.7% and 62.5 ± 13%, respectively in the LF group. The CysCRR was -9.7 ± 6.7% and 29.2 ± 11% in LF and HF hemodialysis, respectively. The statistically significant decrease in CysCRR in the HF group shows the effective clearance of MM by HF dialyzers. Hence, CysCRR could be applied to measure the MM clearance in HF hemodialysis. This study highlights the significance of cystatin C as an important dialysis adequacy marker replacing the conventional markers such as urea and creatinine in HF hemodialysis. Among the middle molecules cystatin C scores over beta-2 microglobulin.

A sensitive WST-8-based bioassay for PEGylated granulocyte colony stimulating factor using the NFS-60 cell line.

CONTEXT: Granulocyte colony stimulating factor (G-CSF) has been commonly used to treat neutropenia caused by chemotherapy, radiotherapy, and organ transplants. Improved in vitro efficacy of G-CSF has already been observed by conjugating it to polyethylene glycol (PEG). OBJECTIVE: The in vivo bioassay using tetrazolium dye with the NFS-60 cell line has been recommended for G-CSF but no such monographs are available for PEGylated G-CSF in pharmacopeias. In the present study, the assay recommended for G-CSF was evaluated for its suitability to PEGylated G-CSF. MATERIALS AND METHODS: The generally used MTS [3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium]-based assay was compared with a bioassay employing a water-soluble tetrazolium dye, WST-8 [2-(2-methoxy-4-nitrophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfophenyl)-2H-tetrazolium], using NFS-60 cells at a concentration of 7 × 10(5) cells/ml against 800 IU/ml of PEGylated G-CSF at 24, 48, 72, and 72 h time points to determine the efficacy of PEGylated G-CSF. Further, the optimized WST-8 dye-based assay was used to test the potency of various commercially available PEGylated G-CSF preparations. RESULTS: The results demonstrated enhanced sensitivity of the WST-8-based assay over the conventional MTS-based assay for determining the potency of PEGylated G-CSF using the NFS-60 cell line. CONCLUSION: Our study demonstrates the potential application of WST-8-based bioassays for other biotherapeutic proteins of human and veterinary interest.

Refolding of recombinant human granulocyte colony stimulating factor: effect of cysteine/cystine redox system.

Granulocyte colony-stimulating factor (G-CSF) is a multifunctional cytokine which is widely used for treating neutropenia in humans. Evaluation of alternative to expensive components of redox buffer (reduced and oxidized glutathione) is an important step in reducing the cost of production of human biotherapeutic proteins. In the present study, refolding of recombinant human G-CSF expressed as inclusion bodies (IBs) in E. coli was optimized using cysteine and cystine redox agents. The refolding to correct native form of G-CSF was assessed by reverse phase high performance liquid chromatography (RP-HPLC). The optimized concentrations of cysteine and cystine for correct refolding of G-CSF were found to be 2 mM and 1 mM, respectively. The correctly refolded G-CSF was detected as early as 4 h of incubation in renaturation buffer containing optimized concentrations of cysteine (2 mM) and cystine (1 mM) redox agents. Refolding of G-CSF in optimized redox system increased with increase in shuffling time. Overall, the results suggested the use of cysteine/cystine redox pair could be an alternative to the costlier redox pairs for successful refolding of G-CSF and possibly other human biotherapeutic proteins of importance.

Pregnancy in a rudimentary uterine horn.

A case of pregnancy reaching term in a rudimentary non-communicating horn of the uterus is presented. The pregnancy resulted in the delivery of a live child by cesarian section. The case is being reported for the rarity with which the foetus survives in this condition.

Amplification and activation of c-Ki-ras oncogene in cell line developed from human pancreas explants.

Amplification and activation of c-Ki-ras gene was studied in normal human pancreas and a cell line (T-3) derived from normal pancreas explants exposed to methylnitrosourea (MNU) for 26 weeks. Normal genomic DNAs from pancreas and derived cell lines showed no transforming activity in NIH 3T3 cells. However, DNAs isolated from tumorigenic cell line derived from MNU treated human pancreas explants transformed NIH 3T3 cells. The hybridization profiles showed that the c-Ki-ras gene was amplified 5 fold in the tumorigenic cells (T-3). The level of mRNA specific to the c-Ki-ras gene was found to be 50-60 fold higher in the malignant cells than in normal human pancreas. These results suggest that higher expression of ras genes is due to gene amplification and/or activation, which is an important step in carcinogenesis.

MNU-induced point mutation and activation of c-Ha-ras oncogene in cell lines derived from human pancreas explants.

Point mutation and activation of c-Ha-ras oncogene was studied at various stages of carcinogenesis in cell lines developed from MNU-treated human pancreas explants. DNAs from normal pancreas and nontumorigenic cell lines showed no transforming activity in NIH 3T3 cells whereas DNA from one of the tumorigenic cell lines transformed NIH 3T3 cells. In this cell line the point mutation was demonstrated to be at codon 12 of c-Ha-ras gene by the loss of an Msp I site. The mutation possibly affected the transcription of c-Ha-ras gene which in turn contributed to the transformation of these cells.

Assembly of the mitochondrial ribosomes in a temperature-conditional mutant of Saccharomyces cerevisiae defective in the synthesis of the var1 protein.

An investigation of the role of the var1 protein in the assembly of the yeast mitochondrial ribosomes was carried out in a temperature conditional mutant, strain h56, which contains a mutation (tsv1) just upstream of the structural gene for the var1 protein. The mutation results in a marked decrease in the synthesis of the var1 protein at the permissive temperature of 28 degrees C and an apparently complete absence of var1 synthesis at the restrictive temperature of 36 degrees C. Long-term growth of strain h56 at the non-permissive temperature was found to result in the loss of the small (37 S) ribosomal subunit and the appearance of a novel 30 S ribonucleoparticle. Both the small (37 S) and the large (54 S) mitochondrial ribosomal subunits were found to be assembled in strain h56 for at least 3 h after transfer to the non-permissive temperature.

Defective assembly of the mitochondrial ribosomes in yeast cells grown in the presence of mitochondrial protein synthesis inhibitors.

The involvement of mitochondrial protein synthesis in the assembly of the mitochondrial ribosomes was investigated by studying the extent to which the assembly process can proceed in the presence of mitochondrial protein synthesis inhibitors erythromycin and chloramphenicol. Yeast cells grown in the presence of erythromycin (2 mg/ml) do not appear to contain any detectable amounts of the mitochondrial small (37 S) ribosomal subunit. Instead, a ribonucleoparticle with a sedimentation coefficient of 30 S was observed; this particle could be shown to be related to the mitochondrial small ribosomal subunit by two-dimensional gel electrophoretic analysis of its protein components. Since the var1 protein is the only mitochondrial translation product known to be associated with the mitochondrial ribosome, our results suggest that this protein is essential for the assembly of the mature small subunit, and that the var1 protein enters the pathway for the assembly of the small subunit at a late step. In at least one strain of yeast the accumulation of the 30-S particle appears to be very sensitive to catabolite repression. When yeast cells are grown in the presence of chloramphenicol instead of erythromycin, assembly of the small subunit appears to be only partially inhibited, and the presence of the 30-S particle could not be clearly demonstrated. This observation is consistent with the fact that in yeast, chloramphenicol inhibits mitochondrial protein synthesis by about 95% only and that the synthesis of the var1 protein appears to be the least sensitive to this inhibition.

The formation of a defective small subunit of the mitochondrial ribosomes in petite mutants of Saccharomyces cerevisiae.

The involvement of mitochondrial protein synthesis in the assembly of the mitochondrial ribosomes was investigated by studying the extent to which the assembly process can proceed in petite mutants of Saccharomyces cerevisiae which lack mitochondrial protein synthetic activity due to the deletion of some tRNA genes and/or one of the rRNA genes on the mtDNA. Petite strains which retain the 15-S rRNA gene can synthesize this rRNA species, but do not contain any detectable amounts of the small mitochondrial ribosomal subunit. Instead, a ribonucleoparticle with a sedimentation coefficient of 30 S (instead of 37 S) was observed. This ribonucleoparticle contained all the small ribosomal subunit proteins with the exception of the var1 and three to five other proteins, which indicates that the 30-S ribonucleoparticle is related to the small mitochondrial ribosomal subunit (37 S). Reconstitution experiments using the 30-S particle and the large mitochondrial ribosomal subunit from a wild-type yeast strain indicate that the 30-S particle is not active in translating the artificial message poly(U). The large mitochondrial ribosomal subunit was present in petite strains retaining the 21-S rRNA gene. The petite 54-S subunit is biologically active in the translation of poly(U) when reconstituted with the small subunit (37 S) from a wild-type strain. The above results indicate that mitochondrial protein synthetic activity is essential for the assembly of the mature small ribosomal subunit, but not for the large subunit. Since the var1 protein is the only mitochondrial translation product known to date to be associated with the mitochondrial ribosomes, the results suggest that this protein is essential for the assembly of the mature small subunit.