Citations

Succinate dehydrogenase (SDH) is a key enzyme in the tricarboxylic acid (TCA) cycle, as well as Complex II of the electron transport chain (ETC). In the SDH reaction, succinate is oxidized to fumarate in the TCA cycle, producing FADH2 contributing to the proton gradient that drives the reduction of ubiquinone to ubiquinol in the ETC. Thus, measurement of the maximum velocity of the SDH reaction (SDHmax) provides insight into both TCA cycle and ETC contributions to the maximum respiratory capacity of mitochondria. The goal of the present study was to use a quantitative histochemical technique, which we previously developed in skeletal muscle fibers, to quantify the SDHmax in permeabilized (45 μg/mL saponin) cardiomyocytes isolated from male Sprague-Dawley rats. In the SDH reaction, the oxidation of succinate (80 mM succinate; determined to be the maximum substrate concentration) to fumarate is coupled to the reduction of nitroblue tetrazolium (NBT; 1.5 mM) to its diformazan (NBTdfz) using 1-methoxyphenazine methosulphate (mPMS; 1 mM) as an exogenous electron carrier and azide (0.75 mM) to inhibit Complex IV. The progressive accumulation of NBTdfz as the reaction proceeds is measured by repeated densitometric (at 570 nm, the peak absorbance wavelength for NBTdfz) measurements. The optical density (OD) measurements are converted to mM fumarate using the Beer-Lambert-Bouger equation based on pathlength for light absorbance and the molar extinction coefficient for NBTdfz. Importantly, the pathlength for light absorbance must be controlled. In cross-sections of skeletal muscle fibers, pathlength was controlled by cryosectioning at 6 μm. In the present study, pathlength was controlled by optical sectioning using a confocal microscope. The isolated cardiomyocytes were imaged in 3D using a Nikon Eclipse A1 laser scanning confocal system with a ×40x/1.3 NA oil-immersion lens with an optical slice set at 0.5 μm. Changes in OD within a series of 10 optical slices were measured every 15 s over a 5 min period and expressed as mM fumarate/min using the Beer-Lambert-Bouger equation. We found that the SDH reaction was linear across 5 min, and that the change in OD was consistent throughout all 10 optical slices within a cardiomyocyte, suggesting that NBTdfz accumulates in the cytoplasm. Our results confirm that this assay of SDHmax is reproducible in cardiomyocytes and offers a single cell alternative to high-resolution respirometry. The ability to measure SDHmax in single cells is in contrast to respirometry, which only measures the average O2 consumption of aggregates of cells (isolated or in tissue). Thus, a direct measurement of SDHmax can provide valuable physiological insight into variation in mitochondrial function across a population of cardiomyocytes, e.g., as might occur with ischemic injury. Supported by the Mayo Clinic This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Progressive multifocal leukoencephalopathy rarely occurs in patients with multiple myeloma. Intracranial central nervous system invasion is also an uncommon event in multiple myeloma, occurring in less than 1% of cases. We describe herein an exceptional case of coexisting progressive multifocal leukoencephalopathy and intraparenchymal central nervous system myeloma infiltration. A 73-year-old woman with relapsed multiple myeloma was treated with 15 cycles of lenalidomide and dexamethasone, but therapy had to be stopped because of a hip fracture after a fall. During hospitalization, the patient developed progressive multifocal leukoencephalopathy caused by John Cunningham virus, and a prominent intra-parenchymal CD138-positive infiltrate was detected. VDJ rearrangements of the immunoglobulin heavy chain gene and the mutational profile of plasma cells in bone marrow at the time of diagnosis and in brain biopsy after progression were analyzed by next generation sequencing, showing genetic differences between medullary and extramedullary myeloma cells. The role of long-term treatment with lenalidomide and dexamethasone in the development progressive multifocal leukoencephalopathy or intraparenchymal central nervous system myeloma infiltration remains unknown. However, our results suggest that both events may have arisen as a consequence of treatment-related immunosuppression. Thus, an appropriate clinical approach compatible with the simultaneous treatment of progressive multifocal leukoencephalopathy and multiple myeloma should be developed.

SummaryCoronary endothelial‐dependent microvascular dysfunction, an early reversible stage of coronary artery disease (CAD), is associated with poor clinical outcome. The current study investigated whether coronary artery distensibility index (CDI) is associated with: (i) coronary endothelial‐dependent microvascular dysfunction and (ii) vulnerable plaque composition among subjects with non‐obstructive CAD. Seventy‐four subjects with non‐obstructive CAD (luminal stenosis <30%) were studied. In 20 subjects with and without coronary endothelial‐dependent microvascular dysfunction, coronary flow reserve (CFR) of target segment during intracoronary (IC) infusion of acetylcholine (Ach) and bolus injection of adenosine as well as CDI at rest of corresponding target segment were measured. In 54 subjects, plaque compositions and CDI at rest of 154 non‐obstructive coronary segments as well as proximal segment without disease were measured by intravascular ultrasound (IVUS). CDI was defined as: [(Early‐diastolic cross‐sectional‐area (CSA) – End‐diastolic CSA of target segment)/(end‐diastolic CSA of target segment × coronary‐pulse‐pressure) × 103]. There is a direct association between endothelial dysfunction and impaired CDI of a coronary segment both in the given coronary segment and corresponding microvessels in which a strong agreement between CDI and CFR Ach (r2 = 0·85, P = 0·0001) was observed. Multivariable regression‐analysis showed that CDI was an independent predictor of the vulnerable plaque characteristics. The risk of impaired CDI was 125% higher in segments with necrotic core and 60% higher in segments with fibrofatty components as compared to normal segments (P = 0·001). In conclusions, the current study reveals that impaired CDI is an endothelial‐dependent process of both given coronary segment and corresponding microvessels and is associated with vulnerable plaque composition.