Production of genetically stable high-titer retroviral vectors that carry a human gamma-globin gene under the control of the alpha-globin locus control region

The ability to generate stable high-titer vectors that give rise to high levels of expression of transduced globin genes in erythroid cells is a prerequisite for effective retroviral-mediated globin gene therapy. The human beta-globin gene with its immediate flanking sequences does not contain all the regulatory elements necessary for regulated high-level and position-independent expression in erythroid cells. The regulatory element known as the beta-globin locus control region (BetaLCR) can provide a linked Beta-globin gene with these properties. However, addition of BetaLCR sequences to a retrovirus carrying a beta-globin gene increases its genetic instability. We have developed a new generation of retroviral vectors in which a human gamma- globin gene is placed under the control of the alphaLCR, the major regulatory element of the alpha-globin gene cluster. We demonstrate that these retroviruses are genetically stable in producer cell lines and can be produced at high titers that exceed 5 x 10(6) colony-forming units (CFU)/mL. In addition, we show that the transduced gamma-globin gene can be expressed in the adult erythroid environment of mouse erythroleukemia (MEL) cells at a level comparable to that of a single endogenous Betamaj-globin gene. These retroviruses can also transduce primary murine bone marrow progenitor cells as efficiently as retroviruses that carry the neomycin resistance (neor) gene. This new generation of globin retroviral vectors may prove useful for gene therapy of human beta-globin gene disorders such as sickle cell disease and beta-thalassemia.     

Synergistic effect of apolipoprotein E epsilon4 and butyrylcholinesterase K-variant on progression from mild cognitive impairment to Alzheimer's disease

OBJECTIVE: To evaluate the synergistic effects of the apolipoprotein E (APOE) epsilon4 and butyrylcholinesterase K-variant (BCHE-K) alleles on progression to Alzheimer's disease (AD) in individuals with mild cognitive impairment (MCI). METHODS: This was a post-hoc exploratory analysis from a 3-4-year, randomized, placebo-controlled study of rivastigmine in participants with MCI (InDDEx study). Participants who consented to genetic testing were included in the current analyses. The incidence of progression to AD, cognitive decline and changes in MRI brain volumes were investigated in participants from the placebo arm of the InDDEx study. RESULTS: Of the 1018 participants in the overall study, 464 were successfully genotyped for both APOE and butyrylcholinesterase. Of these, 68 (14.7%) carried > or =1 APOE epsilon4 and > or =1 BCHE-K allele. The presence of APOE epsilon4 was associated with a significantly higher incidence of progression to AD whereas the presence of BCHE-K had no independent effect on progression. A synergistic effect of the combined presence of APOE epsilon4 and BCHE-K on the time to clinical diagnosis of AD and on MRI brain volumes was seen. Progression to AD and hippocampal volumetric loss was greatest in participants who carried both APOE epsilon4 and BCHE-K alleles and lowest in BCHE-K carriers without the APOE epsilon4 allele. CONCLUSION: In MCI, the risk of cognitive decline, hippocampal volumetric loss and progression to AD seems to be the greatest in individuals who carry at least one copy of both the BCHE-K and APOE epsilon4 alleles.     

The renal toxicity of cadmium metallothionein: Morphometric and X-ray microanalytical studies

This report describes changes in the renal lysosome system of rabbits during a tubular nephropathy produced by a single intravenous injection of cadmium metallothionein at dosages of 0, 0.1, 0.2, or 0.4 mg of Cd/kg body weight. Pronounced light microscopic and ultrastructural changes in renal tubule cells which consisted of necrosis and sloughing were observed at the highest dose. Ultrastructural morphometry and X-ray microanalysis of tubule cells were evaluated in relation to renal function and tissue cadmium levels in the same animals. The most pronounced ultrastructural change in tubule cells of treated animals at all dosages was a highly significant dose-related increase in the volume density of lysosomes which were shown by X-ray microanalysis to contain cadmium. Minor changes were observed in the volume densities of tubule cell mitochondria, nuclei, and vacuoles. The findings were associated with dose-related increases in serum creatinine concentrations and decreases in urine creatinine concentrations. The above data are discussed in relation to tissue concentrations of cadmium and the role of lysosomes in renal tubular nephropathy produced by metal-binding proteins.     

Increased Hepatic and Decreased Urinary Metallothionein in Rats after Cessation of Oral Cadmium Exposure

Abstract: We investigated the role of metallothionein (MT) in tissues after cessation of cadmium (Cd) exposure. Wistar rats of both genders were given CdCl₂ in drinking water at daily doses of 0, 2.5, 5.0 or 10.0 mg Cd/kg body‐weight for 12 weeks. Half of the animals were then killed; the others were given Cd‐free water for the following 16 weeks, i.e. until 28 weeks after start of the experiment (28‐week rats). We observed dose‐dependent increases in the levels of MT in the tissues of rats 12 weeks after beginning the experiment (12‐week rats). After the exposure ceased, levels of MT in the 28‐week rats changed in three ways: an increase in the liver, persistence in the kidney cortex and a decrease in the medulla, relative to those levels in their 12‐week counterparts. Biomarkers of kidney dysfunction were determined to be urinary MT (UMT) and urinary N‐acetyl‐β‐d ‐glucosaminidase (UNAG). After 12 weeks, we observed dose‐related statistically significant increases in UMT and UNAG in all of the Cd‐exposed groups. A statistically significant decrease for UNAG between the 12‐ and 28‐week rats occurred among males at the lowest Cd dose and for UMT in all of the Cd‐exposed groups. The unchanged tissue levels of MT in the kidney cortex suggest that decreased UMT is a sign either of (i) decreased transport of Cd‐MT from the liver via blood plasma to the renal tubules or (ii) increased tubular reabsorption and recovery of renal tubular function.     

The Nordic Trueness Project 2002: use of reference measurement procedure values in a general clinical chemistry survey

Up to 136 laboratories participated in a joint effort to assess the trueness of routine measurements for 14 serum components. An unmodified, fresh-frozen human serum ("IMEP-17 Material 1"), produced for an international inter-laboratory comparison, served as the "master material". The serum had assigned values of the highest available metrological quality, and is assumed to involve no or negligible commutability problems. The material was used in the assignment of traceable values to two other reference sera, "CAL" and "X", through parallel measurements on the three materials according to a common protocol. In this transfer process, uncertainty estimates were provided for all values. The material CAL had been supplied with reference measurement procedure values in 1997, and the two sets of assigned values agreed well. A lyophilized control serum "HK02" was also included in the routine analysis series. It, too, had assigned values based on reference measurement procedures. Significant matrix effects were found. The project has provided: Assigned traceable values for 14 components in a fresh-frozen serum, available to Nordic laboratories for the coming years as "NFKK reference serum X"; Confirmation of earlier assigned reference measurement procedure values for a number of components in CAL, the main calibrator in the Nordic Reference Interval project (NORIP). The transferred values will now serve as the primary reference.; Evidence of long-term stability ( > or = 5 years) of the fresh-frozen serum CAL when stored at -80 degrees C; Evidence of substantial matrix effects in the processed serum HK02. The findings should be used to discuss to what extent reference measurement procedure values are useful and cost-efficient for this type of material.     

Inclusion body myositis in Alzheimer's disease

Roos PM, Vesterberg O, Nordberg M. Inclusion body myositis in Alzheimer’s disease.
Acta Neurol Scand: 2011: 124: 215–217.
© 2010 John Wiley & Sons A/S. Background – The prevalence of Alzheimer’s disease is increasing. Could findings of similar deposits in brain and muscle tissue explain this increase? The purpose of this report is to illustrate that Alzheimer’s disease and inclusion body myositis may share a common aetiology. Results – We present a case where Alzheimer’s disease and inclusion body myositis coexist in the same patient. Amyloid‐beta deposition and the presence of phosphorylated tau protein have been noted in brain tissue and in muscle biopsy from patients with these disorders. Methods – Electrophysiological methods are needed for proper diagnosis of this brain and muscle disorder. Recent data on deposit structures in both conditions may indicate an environmental aetiology for Alzheimer’s disease and inclusion body myositis. Conclusion – By combining electrophysiological methods with muscle biopsy in cases of Alzheimer’s disease, the possible aetiological connection between simultaneous affection of both muscle and brain in this condition can be established.