Mutations in DARS2 result in global dysregulation of mRNA metabolism and splicing.

Leukoencephalopathy with brainstem and spinal cord involvement and lactate elevation (LBSL) is a rare neurological disorder caused by the mutations in the DARS2 gene, which encodes the mitochondrial aspartyl-tRNA synthetase. The objective of this study was to understand the impact of DARS2 mutations on cell processes through evaluation of LBSL patient stem cell derived cerebral organoids and neurons. We generated human cerebral organoids (hCOs) from induced pluripotent stem cells (iPSCs) of seven LBSL patients and three healthy controls using an unguided protocol. Single cells from 70-day-old hCOs were subjected to SMART-seq2 sequencing and bioinformatic analysis to acquire high-resolution gene and transcript expression datasets. Global gene expression analysis demonstrated dysregulation of a number of genes involved in mRNA metabolism and splicing processes within LBSL hCOs. Importantly, there were distinct and divergent gene expression profiles based on the nature of the DARS2 mutation. At the transcript level, pervasive differential transcript usage and differential spliced exon events that are involved in protein translation and metabolism were identified in LBSL hCOs. Single-cell analysis of DARS2 (exon 3) showed that some LBSL cells exclusively express transcripts lacking exon 3, indicating that not all LBSL cells can benefit from the "leaky" nature common to splice site mutations. At the gene- and transcript-level, we uncovered that dysregulated RNA splicing, protein translation and metabolism may underlie at least some of the pathophysiological mechanisms in LBSL. To confirm hCO findings, iPSC-derived neurons (iNs) were generated by overexpressing Neurogenin 2 using lentiviral vector to study neuronal growth, splicing of DARS2 exon 3 and DARS2 protein expression. Live cell imaging revealed neuronal growth defects of LBSL iNs, which was consistent with the finding of downregulated expression of genes related to neuronal differentiation in LBSL hCOs. DARS2 protein was downregulated in iNs compared to iPSCs, caused by increased exclusion of exon 3. The scope and complexity of our data imply that DARS2 is potentially involved in transcription regulation beyond its canonical role of aminoacylation. Nevertheless, our work highlights transcript-level dysregulation as a critical, and relatively unexplored, mechanism linking genetic data with neurodegenerative disorders.

Translational challenges in advancing regenerative therapy for treating neurological disorders using nanotechnology.

The focus of regenerative therapies is to replace or enrich diseased or injured cells and tissue in an attempt to replenish the local environment and function, while slowing or halting further degeneration. Targeting neurological diseases specifically is difficult, due to the complex nature of the central nervous system, including the difficulty of bypassing the brain's natural defense systems. While cell-based regenerative therapies show promise in select tissues, preclinical and clinical studies have been largely unable to transfer these successes to the brain. Advancements in nanotechnologies have provided new methods of central nervous system access, drug and cell delivery, as well as new systems of cell maintenance and support that may bridge the gap between regenerative therapies and the brain. In this review, we discuss current regenerative therapies for neurological diseases, nanotechnology as nanocarriers, and the technical, manufacturing, and regulatory challenges that arise from inception to formulation of nanoparticle-regenerative therapies.

Neural effects of inflammation, cardiovascular disease, and HIV: Parallel, perpendicular, or progressive?

The pervasive reach of the inflammatory system is evidenced by its involvement in numerous disease states. Cardiovascular disease, marked by high levels of circulating inflammatory mediators, affects an estimated 83.6 million Americans. Similarly, human immunodeficiency virus (HIV) produces a paradoxical state of generalized immune activity despite widespread immunosuppression, and affects 35 million people worldwide. Patients living with HIV (PLWH) suffer from inflammatory conditions, including cardiovascular disease (CVD), at a rate exceeding the general population. In this combined disease state, immune mechanisms that are common to both CVD and HIV may interact to generate a progressive condition that contributes to the exacerbated pathogenesis of the other to the net effect of damage to the brain. In this review, we will outline inflammatory cell mediators that promote cardiovascular risk factors and disease initiation and detail how HIV-related proteins may accelerate this process. Finally, we examine the extent to which these comorbid conditions act as parallel, perpendicular, or progressive sequela of events to generate a neurodegenerative environment, and consider potential strategies that can be implemented to reduce the burden of CVD and inflammation in PLWH.

Neoadjuvant bevacizumab persistently inactivates VEGF at the time of surgery despite preoperative cessation.

BACKGROUND: When anti-VEGF (vascular endothelial growth factor) antibody bevacizumab is applied in neoadjuvant treatment of colorectal cancer patients with liver metastasis, 5-6 weeks between last bevacizumab dose and liver resection are currently recommended to avoid complications in wound and liver regeneration. In this context, we aimed to determine whether VEGF is inactivated by bevacizumab at the time of surgery. METHODS: Fifty colorectal cancer patients with liver metastases received neoadjuvant chemotherapy ± bevacizumab supplementation. The last dose of bevacizumab was administered 6 weeks before surgery. Plasma, subcutaneous and intraabdominal wound fluid were analysed for VEGF content before and after liver resection (day 1-3). Immunoprecipitation was applied to determine the amount of bevacizumab-bound VEGF. RESULTS: Bevacizumab-treated individuals showed no increase in perioperative complications. During the entire monitoring period, plasma VEGF was inactivated by bevacizumab. In wound fluid, VEGF was also completely bound by bevacizumab and was remarkably low compared with the control chemotherapy group. CONCLUSION: These data document that following a cessation time of 6 weeks, bevacizumab is fully active and blocks circulating and local VEGF at the time of liver resection. However, despite effective VEGF inactivation no increase in perioperative morbidity is recorded suggesting that VEGF activity is not essential in the immediate postoperative recovery period.

Decreased expression of the melatonin receptor 1 in human colorectal adenocarcinomas.

Melatonin exerts anti-proliferative and pro-apoptotic effects in various cancer cell lines. Furthermore, there is evidence for impaired melatonin secretion in human breast and colorectal cancer. Additionally, several studies revealed a modulated expression of the melatonin receptor 1 (MT1), in human breast cancer specimens. Since melatonin binding sites were already identified in the human intestine, our aim is to identify the expression and to characterize the localization of the MT1 receptor in the human colon and in particular to compare MT1 expression levels between non-malignant and malignant colonic tissue. We assessed MT1 transcript levels with real time RT-PCR in colon adenocarcinomas and the adjacent normal colonic mucosa of 39 patients and observed a significant decrease of MT1 mRNA expression in colorectal cancer compared with the healthy adjacent mucosa tissue (0.67 mean difference, P < 0.0001). The results were confirmed at the protein level by Western blot analysis and by immunohistochemistry. MT1 was localized mainly supranuclear in colonic epithelial cells lining the crypts. We also evaluated mRNA expression of different clock genes in the colon samples and found a significant correlation between MT1 and Cryptochrome 1 (Cry1) expression (P < 0.01 for normal and P < 0.05 for tumour tissue). In conclusion, the decreased expression of MT1 in human colorectal cancer could point to a role of melatonin in this disease.

Temporal and spatial changes in cell wall composition in developing grains of wheat cv. Hereward.

A combination of enzyme mapping, FT-IR microscopy and NMR spectroscopy was used to study temporal and spatial aspects of endosperm cell wall synthesis and deposition in developing grain of bread wheat cv. Hereward. This confirmed previous reports that changes in the proportions of the two major groups of cell wall polysaccharides occur, with beta-glucan accumulating earlier in development than arabinoxylan. Changes in the structure of the arabinoxylan occurred, with decreased proportions of disubstituted xylose residues and increased proportions of monosubstituted xylose residues. These are likely to result, at least in part, from arabinoxylan restructuring catalysed by enzymes such as arabinoxylan arabinofurano hydrolase and lead to changes in cell wall mechanical properties which may be required to withstand stresses during grain maturation and desiccation.

Thrombospondin-1: a unique marker to identify in vitro platelet activation when monitoring in vivo processes.

BACKGROUND: Measuring platelet activation in patients has become a potent method to investigate pathophysiological processes. However, the commonly applied markers are sensitive to detrimental influences by in vitro platelet activation during blood analysis. OBJECTIVES: Protein isoforms of platelet-derived thrombospondin-1 (TSP-1) were investigated for their potential to identify in vitro platelet activation when monitoring in vivo processes. METHODS: TSP-1 was determined in plasma, serum or supernatant of purified platelets by ELISA and immunoblotting and was compared with standard markers of platelet activation. A collective of 20 healthy individuals and 30 cancer patients was analyzed. RESULTS: While in vitro platelet degranulation led to a selective increase in the 200-kDa full-length molecule, an in vivo process involving platelet activation such as wound healing resulted in the predominant rise of the 140-kDa TSP-1 protein. The physiological ratio of circulating TSP-1 variants was determined and a cut-off level at 1.0 was defined to identify plasma samples with artificial in vitro platelet activation exceeding the cut-off level. In contrast, cancer patients known to frequently exhibit increased in vivo activation of platelets presented with a significantly decreased ratio of TSP-1 variants as compared with healthy volunteers. CONCLUSIONS: In comparison to standard platelet markers, TSP-1 constitutes a sensitive and stable parameter suited to monitor in vitro platelet activation. The analysis of TSP-1 protein isoforms further offers a valuable tool to reliably discriminate between in vitro and in vivo effects, to exclude variability introduced during blood processing and improve clinical monitoring.

Radiation dose from coal slag used as building material in the Transdanubian region of Hungary.

Coals mined in the Transdanubian region in Hungary have an elevated concentration of (226)Ra, which becomes enriched in the slag after burning. This slag has been used as filling and/or insulating material in building works. The aim of this study was to investigate the radiological situation in this territory in terms of the possible impact of this residual material from coal. Flats in three towns with a coal mine and a coal-fired power plant operating in their neighbourhood were examined. The radionuclide contents (including (226)Ra, (232)Th and (40)K) of the slag used for building were determined, and the slags were categorised according to the international standards and recommendations. The external gamma dose rate and the radon concentration in the sites were measured, and based on these data dose assessments were made. The (226)Ra concentration of the slag was 160-2,893 Bq kg(-1); the indoor gamma dose rates were 82-633 nGy h(-1); the radon concentration measured with a nuclear track detector varied from 29 to 1,310 Bq m(-3); the assessed dose contributions in the three towns were 0.65-1.57 mSv y(-1) due to gamma radiation and 2.2-15.2 mSv y(-1) due to radon.

Getting to the point: developing IT for the sharp end of healthcare.

Healthcare demonstrates the same properties of risk, complexity, uncertainty, dynamic change, and time-pressure as other high hazard sectors including aviation, nuclear power generation, the military, and transportation. Unlike those sectors, healthcare has particular traits that make it unique such as wide variability, ad hoc configuration, evanescence, resource constraints, and governmental and professional regulation. While healthcare's blunt (management) end is more easily understood, the sharp (operator) end is more difficult to research the closer one gets to the sharp end's point. Understanding sharp end practice and cognitive work can improve computer-based systems resilience, which is the ability to perform despite change and challenges. Research into actual practice at the sharp end of healthcare will provide the basis to understand how IT can support clinical practice. That understanding can be used to develop computer-based systems that will act as team players, able to support both individual and distributed cognitive work at healthcare's sharp end.

Assessing risk: the role of probabilistic risk assessment (PRA) in patient safety improvement.

Morbidity and mortality due to "medical errors" compel better understanding of health care as a system. Probabilistic risk assessment (PRA) has been used to assess the designs of high hazard, low risk systems such as commercial nuclear power plants and chemical manufacturing plants and is now being studied for its potential in the improvement of patient safety. PRA examines events that contribute to adverse outcomes through the use of event tree analysis and determines the likelihood of event occurrence through fault tree analysis. It complements tools already in use in patient safety such as failure modes and effects analyses (FMEAs) and root cause analyses (RCAs). PRA improves on RCA by taking account of the more complex causal interrelationships that are typical in health care. It also enables the analyst to examine potential solution effectiveness by direct graphical representations. However, PRA simplifies real world complexity by forcing binary conditions on events, and it lacks adequate probability data (although recent developments help to overcome these limitations). Its reliance on expert assessment calls for deep domain knowledge which has to come from research performed at the "sharp end" of acute care.

Diurnal blood pressure variations in incipient and end stage diabetic renal disease.

Our aim was to compare the diurnal blood pressure patterns of people with Type 1 diabetes on continuous ambulatory peritoneal dialysis (CAPD, n=9) or haemodialysis (n=10) to diabetic patients with normo-albuminuria (n=12) or micro-albuminuria (n=15). Blood pressure was measured with an ABPM02 Meditech oscillometric blood pressure monitor. The micro-albuminuric group had significantly higher nocturnal diastolic and mean arterial pressures than the normo-albuminuric group. CAPD and haemodialysis patients had significantly higher day time, nocturnal mean systolic, diastolic and mean arterial blood pressures. Micro-albuminuric and end-stage renal failure patients displayed a loss of the physiological drop of systolic blood pressure, which was only significant in the normo-albuminuric group. Nocturnal drop of blood pressure characterised by diurnal indices were 7.4% in the CAPD, 8.8% in the haemodialysis, 10.0% in the micro-albuminuric and 16.5% in the normo-albuminuric group. These results suggest, that pathological circadian blood pressure variation is common in diabetic patients on dialysis, and ambulatory blood pressure monitoring can be a useful tool both in its the detection and its adequate treatment.

Radiation hazard of coal-slags as building material in Tatabánya town (Hungary).

High concentrations of 226Ra (865-2,383 Bq kg(1)) were measured in the coal-slags, originated from the region of the settlement Tatabánya, Transdanubian Middle Mountains, Hungary. These slags are commonly used as building materials in this district. The external gamma dose rate was measured in 188 rooms at different heights above the floor. In 124 rooms with slags used for construction, the average absorbed dose rate was 296 nGy h(-1). In 10 apartments the average radon concentration was 502 Bq m(-3). In that case the estimated effective dose due to inhaled radon and its progeny and gamma radiation was 10.3 mSv y(-1).

Diurnal blood pressure variation and albuminuria in normotensive patients with insulin-dependent diabetes mellitus.

BACKGROUND: Abnormalities of the systemic blood pressure are closely associated with the development of diabetic nephropathy. Our aim was to examine the relationship between diurnal blood pressure pattern and albuminuria in insulin-dependent normotensive diabetic patients before the development of overt nephropathy. METHODS: Urinary albumin excretion rates were determined by radioimmunoassay, and 24-h ambulatory blood pressure monitoring was performed. Means and diurnal index was calculated for systolic, diastolic and mean arterial blood pressure, for day-time, night-time, and the whole day. The results of the normoalbuminuric (n = 39) and microalbuminuric (n = 29) groups are compared, and correlation of the blood pressure parameters with albuminuria is analysed. RESULTS: Twenty-four hours and night-time mean blood pressures were significantly higher, diurnal indices characterizing the night-time blood pressure drop were smaller in the microalbuminuric group. With multiple regression analysis a significant positive correlation was found between albumin excretion rates and 24-h mean systolic blood pressure and a significant negative correlation between albumin excretion rates and the diurnal index of mean arterial pressure (r2= 0.40, P<0.0001). In the normoalbuminuric group 1 (2.6%) patient, in the microalbuminuric group 7 (24.1%/) were 'non-dippers'. CONCLUSION: We conclude that in normotensive insulin-dependent diabetic patients the night-time decrease of blood pressure is smaller if microalbuminuria is present. Higher nocturnal blood pressure load is associated with the increase of albuminuria, even before the onset of overt diabetic nephropathy or hypertension.

The efficacy of long-term captopril treatment on micro- and macroalbuminuria in hypertensive diabetics.

Microalbuminuric [16] and macroalbuminuric [17] hypertensive insulin dependent diabetics were followed up for 4 years after the initiation of captopril therapy to assess the efficacy of ACE inhibitor therapy on albuminuria and blood pressure normalisation. Within the first six months of captopril therapy mean systolic blood pressure decreased in microalbuminuric and macroalbuminuric patients from 168.1 +/- 17.6 mmHg to 134.4 +/- 12.1 mmHg (19.2 +/- 7.1%) and from 177.6 +/- 16.8 mmHg to 143.5 +/- 12.7 (18.9 +/- 6.7%) mmHg, respectively. Mean diastolic blood pressure, similarly, showed a decrease from 91.9 +/- 9.1 mmHg to 74.4 +/- 10.3 mmHg (19.0 +/- 9.4%) in the microalbuminuric and from 95.3 +/- 13.7 mmHg to 78.2 +/- 7.3 (16.9 +/- 9.5%) mmHg in the macroalbuminuric group. After six months of captopril administration albumin excretion rates decreased as well, from 97.4 +/- 35.9 micrograms/min to 51.9 +/- 19.9 micrograms/min (46.9 +/- 7.6%) and from 766.7 +/- 577.9 micrograms/min to 365.1 +/- 298.4 micrograms/min (50.4 +/- 8.4%) in the micro- and macroalbuminuric groups, respectively. Thereafter, mean albumin excretion rates and blood pressure rose significantly, but at the end of the fourth year they were still significantly lower compared to that of the pretreatment period. After four years, albumin excretion rates were 71.3 +/- 29.6 micrograms/min in the microalbuminuric and 391.2 +/- 204.7 micrograms/min in the macroalbuminuric group. We conclude that ACE inhibitor therapy results in a rapid decrease of albuminuria and blood pressure, and despite a slow gradual increase, the albumin excretion rates and blood pressure values remain significantly lower than the initial values after four years.

Demystifying AIDS in Thailand: a dialectical analysis of the Thai sex industry.

The AIDS pandemic throughout the world, and particularly in Thailand, is not just a physical disease that has evaded much of our control. It is symptomatic of a chronic, more pervasive and sinister plague that governs modern societies. This plague is social and political in nature. In the case of Thailand's AIDS pandemic, this paper argues that the underlying sociopolitical plague that further aggravates the AIDS crisis may be found in the internal contradictions that govern the Thai sex industry and its comodification of women and sex. It further suggests that current Thai management of the AIDS pandemic, though effective in terms of increasing the public's awareness about AIDS, its transmission modes and preventive measures, is less effective in the long run, because it fails to adequately address structural formations, contradictions, and practices that constitute and reproduce the Thai sex industry. Two objectives are attempted in this paper: first, a dialectical analysis of the internal contradictions in Thai society, as it relates to the sex industry, the family, and Buddhist institutions in Thailand; and, second, a line of response to the AIDS pandemic in Thailand.