Mitochondrial respiration defects modulate differentiation but not proliferation of hematopoietic stem and progenitor cells

Mitochondrial energy production is involved in various cellular processes. Here we show that ATP content is significantly increased in lineage-restricted progenitor cells compared with hematopoietic stem and progenitor cells (HSPCs) or more differentiated cells. Transplantation analysis using a mouse model of mitochondrial disease revealed that mitochondrial respiration defects resulted in a significant decrease in the total number and repopulating activity of bone marrow cells, although the number of HSPCs increased. The proliferative activity of HSPCs and lineage-restricted progenitor cells was not impaired by reduction of ATP content and there seems to be no associated increase in reactive oxygen species levels and apoptosis. Our findings indicate that mitochondrial respiration defects modulate HSPC commitment/differentiation into lineage-restricted progenitor cells.     

The role of mitochondrial transition pores on bovine oocyte competence after heat stress, as determined by effects of cyclosporin A

This study was undertaken to investigate the effects on the nuclear maturation and subsequent embryonic development of bovine oocytes exposed to heat stress (HS) when treating bovine oocytes before in vitro maturation (IVM) with 1 μM cyclosporin A (CsA), an inhibitor of mitochondrial permeability transition pore opening. Mitochondrial activity, reactive oxygen species (ROS), and apoptosis levels of the oocytes were also assessed. Nuclear maturation rates of both the HS-exposed oocytes treated with or without CsA groups (HS + CsA or HS group) were significantly lower (P<0.05) than that of the control group, while the rate of the HS + CsA group was significantly higher (P<0.05) than that of the HS group. Furthermore, although the cleavage and blastocyst formation rates of the HS group were significantly lower than those of the control groups (P<0.05), both rates of the HS + CsA group recovered to the same level as those of the control group. The HS group showed a significantly higher ROS level, lower mitochondrial activity in the oocytes, and TUNEL-positive cumulus cells, but not oocytes, compared with those of the control group (P<0.05), whereas the TUNEL-positive and mitochondrial activity levels of the HS + CsA group recovered to those of the control group. These results indicate that 1 μM CsA treatment before IVM may mitigate reduced mitochondrial activity, increase number of apoptotic cumulus cells under HS, and improve the nuclear maturation and developmental competence of bovine oocytes.     

Stability of DNA/Td3717 complexes for gene transfection,defined by the size and polydispersity of the complex

The amphiphilic α-helical peptide, Td3717, is a bi-functional synthetic peptide that acts as both a polycation for DNA binding and a ligand for targeted delivery to tumor cells. Td3717 forms a stable complex with plasmid DNA, and the complex maintained high transfection efficiency after storage at 4 °C for six months and after four freeze/thaw cycles. During the storage and freeze/thaw cycling, the particle size of the DNA/Td3717 complex remained less than 100 nm. The size of the complex is an important factor for its internalization into cells via the endocytosis pathway; therefore, the stability of the particles will strongly contribute to high transfection efficiencies after storage and repeated freezing/thawing.     

Radioiodine (131I) therapy of Graves' disease: use of the new high resolutional ultrasonic scanner for the determination of the accurate weight of the thyroid gland

In this series, eighteen patients with Graves' disease were treated with 8000 rads (80 Gy) of radioiodine (131I), using the new high resolutional ultrasonic scanner for the determination of the accurate weight of the thyroid gland. The mean dose of radioiodine administered orally was 4.6 +/- 3.0 mCi (170.2 +/- 110.0 MBq) and 133.7 +/- 44.6 microCi/g (4.95 +/- 1.65 MBq). At one year after treatment, twelve of eighteen patients (66.7%) became euthyroid, five (27.8%) remained hyperthyroid and one (5.6%) became hypothyroid. Analysis of various factors which may be related to the effect of radioiodine therapy revealed that the weight of the thyroid gland in the hyperthyroid and euthyroid groups was significantly different (61.7 +/- 33.5 g vs. 25.1 +/- 9.1 g, p less than 0.05). Furthermore, all patients with larger glands (more than sixty grams) remained hyperthyroid, while the incidence of euthyroidism was as high as 80% in patients with smaller glands (less than forty grams). Although the number of patients studied was small, these results indicate that a larger thyroid gland requires a larger radioiodine dose per gram of tissue than a smaller gland, suggesting that the therapeutic radiation dose should be graded according to the gland size even when the gland size is accurately estimated by ultrasound. Further study is required to determine the appropriate radiation dose graded according to the gland size.     

Hepatitis C Related Chronic Liver Cirrhosis: Feasibility of Texture Analysis of MR Images for Classification of Fibrosis Stage and Necroinflammatory Activity Grade

Purpose

To assess the feasibility of texture analysis for classifying fibrosis stage and necroinflammatory activity grade in patients with chronic hepatitis C on T2-weighted (T2W), T1-weighted (T1W) and Gd-EOB-DTPA-enhanced hepatocyte-phase (EOB-HP) imaging.

Materials and methods

From April 2008 to June 2012, MR images from 123 patients with pathologically proven chronic hepatitis C were retrospectively analyzed. Texture parameters derived from histogram, gradient, run-length matrix, co-occurrence matrix, autoregressive model and wavelet transform methods were estimated with imaging software. Fisher, probability of classification error and average correlation, and mutual information coefficients were used to extract subsets of optimized texture features. Linear discriminant analysis in combination with 1-nearest neighbor classifier (LDA/1-NN) was used for lesion classification. In compliance with the software requirement, classification was performed based on datasets from all patients, the patient group with necroinflammatory activity grade 1, and that with fibrosis stage 4, respectively.

Results

Based on all patient dataset, LDA/1-NN produced misclassification rates of 28.46%, 35.77% and 20.33% for fibrosis staging and 34.15%, 25.20% and 28.46% for necroinflammatory activity grading in T2W, T1W and EOB-HP images. In the patient group with necroinflammatory activity grade 1, LDA/1-NN yielded misclassification rates of 5.00%, 0% and 12.50% for fibrosis staging in T2W, T1W and EOB-HP images respectively. In the patient group with fibrosis stage 4, LDA/1-NN yielded misclassification rates of 5.88%, 12.94% and 11.76% for necroinflammatory activity grading in T2W, T1W and EOB-HP images respectively.

Conclusion

Texture quantitative parameters of MR images facilitate classification of the fibrosis stage as well as necroinflammatory activity grade in chronic hepatitis C, especially after categorizing the input dataset according to the activity or fibrosis degree in order to remove the interference between the fibrosis stage and necroinflammatory activity grade on texture features.     

Utility of NucleoCounter for the chondrocyte count in the collagenase digest of human native cartilage

In cartilage tissue engineering, viable cell numbers should be correctly counted in the collagenase digest of the biopsied cartilage. However, this is a difficult task due to the presence of matrix debris, cell ghosts and their aggregates. To search for the correct cell counting method in this situation, we evaluated the utility of an automatic cell counting device, the NucleoCounter, and compared it with conventional staining using the LIVE/DEAD® kit. We first measured the cell numbers of a standard chondrocyte sample by the NucleoCounter, which showed a high accuracy (R² = 0.9999) and reproducibility (%CV: 2.00–8.66). We then calculated the cell numbers and viability in some collagenase digests of native cartilage using either the NucleoCounter or LIVE/DEAD® kit, revealing that the total cell numbers, viable ones and viability were highly correlated between them (R² = 0.9601, 0.9638 and 0.917, respectively). However, both the intrapersonal and interpersonal variabilities in the NucleoCounter was significantly decreased to about 1/20–1/5, compared to that of the LIVE/DEAD® kit. The NucleoCounter was regarded as a useful tool for simple, rapid, and highly reproducible cell counts, which may not only provide constant experimental data in a certain laboratory, but also contribute to the high reproducibility of the clinical results of cartilage tissue engineering among multiple institutions.     

Pathogenic mitochondrial DNA-induced respiration defects in hematopoietic cells result in anemia by suppressing erythroid differentiation

Anemia is a symptom in patients with Pearson syndrome caused by the accumulation of mutated mitochondrial DNA (mtDNA). Such mutated mtDNAs have been detected in patients with anemia. This suggested that respiration defects due to mutated mtDNA are responsible for the anemia. However, there has been no convincing experimental evidence to confirm the pathophysiological relation between respiration defects in hematopoietic cells and expression of anemia. We address this issue by transplanting bone marrow cells carrying pathogenic mtDNA with a large-scale deletion (ΔmtDNA) into normal mice. The bone marrow-transplanted mice carried high proportion of ΔmtDNA only in hematopoietic cells, and resultant the mice suffered from macrocytic anemia. They show abnormalities of erythroid differentiation and weak erythropoietic response to a stressful condition. These observations suggest that hematopoietic cell-specific respiration defects caused by mtDNAs with pathogenic mutations are responsible for anemia by inducing abnormalities in erythropoiesis.