Acquired hypofibrinogenemia in a patient with multiple myeloma.

We report a case of acquired hypofibrinogenemia with multiple myeloma presenting λ-type IgG monoclonal protein. The patient had anemia and renal deficiency, and also developed bleeding tendency due to severe coagulopathy. Her fibrinogen level was under the detectable limits in a functional assay. Enzyme-linked immunosorbent assay (ELISA) and immunoblotting analysis results were consistent with functional assay results, and deficiency patterns observed in cross-mixing tests for PT and aPTT confirmed the diagnosis of hypofibrinogenemia. To determine the cause of hypofibrinogenemia, we purified the patient's immunoglobulin via protein A agarose, and confirmed that fibrinogen was included in the bound fraction, strongly indicating paraprotein interference with fibrinogen. As accelerated removal of fibrinogen was indicated, we incubated the patient's plasma up to 48 h, but did not observe significant loss of fibrinogen. In sharp contrast, fibrinogen returned to below the detection level 12 h after infusion of fresh frozen plasma. These findings support leukocyte-mediated fibrinogen removal, rather than paraprotein-triggered fibrinogen instability. Surprisingly, the patient's paraprotein was IgG2, but we speculate the amount of paraprotein (IgG 5346 mg/dL) compensated for lower affinity to Fcγ receptors.

Homeostasis of the ER redox state subsequent to proteasome inhibition.

Endoplasmic reticulum (ER) maintains within, an oxidative redox state suitable for disulfide bond formation. We monitored the ER redox dynamics subsequent to proteasome inhibition using an ER redox probe ERroGFP S4. Proteasomal inhibition initially led to oxidation of the ER, but gradually the normal redox state was recovered that further led to a reductive state. These events were found to be concomitant with the increase in the both oxidized and reduced glutathione in the microsomal fraction, with a decrease of total intracellular glutathione. The ER reduction was suppressed by pretreatment of a glutathione synthesis inhibitor or by knockdown of ATF4, which induces glutathione-related genes. These results suggested cellular adaptation of ER redox homeostasis: (1) inhibition of proteasome led to accumulation of misfolded proteins and oxidative state in the ER, and (2) the oxidative ER was then reduced by ATF4 activation, followed by influx of glutathione into the ER.

Synthesized Aß42 Caused Intracellular Oxidative Damage, Leading to Cell Death, via Lysosome Rupture.

Neuronal cellular accumulation of amyloid beta peptide (Aß) has been implicated in the pathogenesis of Alzheimer's disease (AD). Intracellular accumulation of Aß42, a toxic form of Aß, was observed as an early event in AD patients. However, its contribution and the cellular mechanism of cell death remained unclear. We herein revealed the mechanism by which Aß42 incorporated into cells leads to cell death by using chemically synthesized Aß42 variants. The Aß42 variant Aß42 (E22P) which has an increased tendency to oligomerize, accumulated in lysosomes at an earlier stage than wild-type Aß42, leading to higher ROS production and lysosomal membrane oxidation, and resulting in cell death. On the other hand, Aß42 (E22V), which is incapable of oligomerization, did not accumulate in cells or affect the cell viability. Moreover, intracellular localization of EGFP-Galectin-3, a ß-galactoside binding lectin, showed that accumulation of oligomerized Aß42 in lysosomes caused lysosomal membrane permeabilization (LMP). Overexpression of lysosome-localized LAMP1-fused peroxiredoxin 1 and treatment with U18866A, an inhibitor of cholesterol export from lysosomes that causes an increase in lysosomal membrane stability, attenuated Aß42-mediated LMP and cell death. Our findings show that lysosomal ROS generation by toxic conformer of Aß led to cell death via LMP, and suggest that these events are potential targets for AD prevention.Key words: Amyloid-beta (Aß), Cell death, Lysosome, Lysosomal membrane permeabilization, Reactive oxygen species (ROS).