Phosphorylation in Novel Mitochondrial Creatine Kinase Tyrosine Residues Render Cardioprotection against Hypoxia/Reoxygenation Injury

Objective@#Ischemic cardiomyopathy (ICM) is the leading cause of heart failure. Proteomic and genomic studies have demonstrated ischemic preconditioning (IPC) can assert cardioprotection against ICM through mitochondrial function regulation. Considering IPC is conducted in a relatively brief period, regulation of protein expression also occurs very rapidly, highlighting the importance of protein function modulation by post-translational modifications. This study aimed to identify and analyze novel phosphorylated mitochondrial proteins that can be harnessed for therapeutic strategies for preventing ischemia/reperfusion (I/R) injury. @*Methods@#Sprague-Dawley rat hearts were used in an ex vivo Langendorff system to simulate normal perfusion, I/R, and IPC condition, after which the samples were prepared for phosphoproteomic analysis. Employing human cardiomyocyte AC16 cells, we investigated the cardioprotective role of CKMT2 through overexpression and how site-directed mutagenesis of putative CKMT2 phosphorylation sites (Y159A, Y255A, and Y368A) can affect cardioprotection by measuring CKMT2 protein activity, mitochondrial function and protein expression changes. @*Results@#The phosphoproteomic analysis revealed dephosphorylation of mitochondrial creatine kinase (CKMT2) during ischemia and I/R, while preserving its phosphorylated state during IPC. CKMT2 overexpression conferred cardioprotection against hypoxia/reoxygenation (H/R) by increasing cell viability and mitochondrial adenosine triphosphate level, preserving mitochondrial membrane potential, and reduced reactive oxygen species (ROS) generation, while phosphomutations, especially in Y368, nullified cardioprotection by significantly reducing cell viability and increasing ROS production during H/R. CKMT2 overexpression increased mitochondrial function by mediating the proliferator-activated receptor γ coactivator-1α/ estrogen-related receptor-α pathway, and these effects were mostly inhibited by Y368A mutation. @*Conclusion@#These results suggest that regulation of quantitative expression and phosphorylation site Y368 of CKMT2 offers a unique mechanism in future ICM therapeutics.

Far-infrared rays enhance mitochondrial biogenesis and GLUT3 expression under low glucose conditions in rat skeletal muscle cells

Far-infrared rays (FIR) are known to have various effects on atoms and molecular structures within cells owing to their radiation and vibration frequencies. The present study examined the effects of FIR on gene expression related to glucose transport through microarray analysis in rat skeletal muscle cells, as well as on mitochondrial biogenesis, at high and low glucose conditions. FIR were emitted from a bio-active material coated fabric (BMCF). L6 cells were treated with 30% BMCF for 24 h in medium containing 25 or 5.5 mM glucose, and changes in the expression of glucose transporter genes were determined. The expression of GLUT3 (Slc2a3) increased 2.0-fold (p < 0.05) under 5.5 mM glucose and 30% BMCF. In addition, mitochondrial oxygen consumption and membrane potential (ΔΨm) increased 1.5- and 3.4-fold (p < 0.05 and p < 0.001), respectively, but no significant change in expression of Pgc-1a, a regulator of mitochondrial biogenesis, was observed in 24 h. To analyze the relationship between GLUT3 expression and mitochondrial biogenesis under FIR, GLUT3 was down-modulated by siRNA for 72 h. As a result, the ΔΨm of the GLUT3 siRNA-treated cells increased 3.0-fold (p < 0.001), whereas that of the control group increased 4.6-fold (p < 0.001). Moreover, Pgc-1a expression increased upon 30% BMCF treatment for 72 h; an effect that was more pronounced in the presence of GLUT3. These results suggest that FIR may hold therapeutic potential for improving glucose metabolism and mitochondrial function in metabolic diseases associated with insufficient glucose supply, such as type 2 diabetes.

Phosphorylation in Novel Mitochondrial Creatine Kinase Tyrosine Residues Render Cardioprotection against Hypoxia/Reoxygenation Injury

Objective@#Ischemic cardiomyopathy (ICM) is the leading cause of heart failure. Proteomic and genomic studies have demonstrated ischemic preconditioning (IPC) can assert cardioprotection against ICM through mitochondrial function regulation. Considering IPC is conducted in a relatively brief period, regulation of protein expression also occurs very rapidly, highlighting the importance of protein function modulation by post-translational modifications. This study aimed to identify and analyze novel phosphorylated mitochondrial proteins that can be harnessed for therapeutic strategies for preventing ischemia/reperfusion (I/R) injury. @*Methods@#Sprague-Dawley rat hearts were used in an ex vivo Langendorff system to simulate normal perfusion, I/R, and IPC condition, after which the samples were prepared for phosphoproteomic analysis. Employing human cardiomyocyte AC16 cells, we investigated the cardioprotective role of CKMT2 through overexpression and how site-directed mutagenesis of putative CKMT2 phosphorylation sites (Y159A, Y255A, and Y368A) can affect cardioprotection by measuring CKMT2 protein activity, mitochondrial function and protein expression changes. @*Results@#The phosphoproteomic analysis revealed dephosphorylation of mitochondrial creatine kinase (CKMT2) during ischemia and I/R, while preserving its phosphorylated state during IPC. CKMT2 overexpression conferred cardioprotection against hypoxia/reoxygenation (H/R) by increasing cell viability and mitochondrial adenosine triphosphate level, preserving mitochondrial membrane potential, and reduced reactive oxygen species (ROS) generation, while phosphomutations, especially in Y368, nullified cardioprotection by significantly reducing cell viability and increasing ROS production during H/R. CKMT2 overexpression increased mitochondrial function by mediating the proliferator-activated receptor γ coactivator-1α/ estrogen-related receptor-α pathway, and these effects were mostly inhibited by Y368A mutation. @*Conclusion@#These results suggest that regulation of quantitative expression and phosphorylation site Y368 of CKMT2 offers a unique mechanism in future ICM therapeutics.

Far-infrared rays enhance mitochondrial biogenesis and GLUT3 expression under low glucose conditions in rat skeletal muscle cells

Far-infrared rays (FIR) are known to have various effects on atoms and molecular structures within cells owing to their radiation and vibration frequencies. The present study examined the effects of FIR on gene expression related to glucose transport through microarray analysis in rat skeletal muscle cells, as well as on mitochondrial biogenesis, at high and low glucose conditions. FIR were emitted from a bio-active material coated fabric (BMCF). L6 cells were treated with 30% BMCF for 24 h in medium containing 25 or 5.5 mM glucose, and changes in the expression of glucose transporter genes were determined. The expression of GLUT3 (Slc2a3) increased 2.0-fold (p < 0.05) under 5.5 mM glucose and 30% BMCF. In addition, mitochondrial oxygen consumption and membrane potential (ΔΨm) increased 1.5- and 3.4-fold (p < 0.05 and p < 0.001), respectively, but no significant change in expression of Pgc-1a, a regulator of mitochondrial biogenesis, was observed in 24 h. To analyze the relationship between GLUT3 expression and mitochondrial biogenesis under FIR, GLUT3 was down-modulated by siRNA for 72 h. As a result, the ΔΨm of the GLUT3 siRNA-treated cells increased 3.0-fold (p < 0.001), whereas that of the control group increased 4.6-fold (p < 0.001). Moreover, Pgc-1a expression increased upon 30% BMCF treatment for 72 h; an effect that was more pronounced in the presence of GLUT3. These results suggest that FIR may hold therapeutic potential for improving glucose metabolism and mitochondrial function in metabolic diseases associated with insufficient glucose supply, such as type 2 diabetes.

Time-dependent proteomic and genomic alterations in Toll-like receptor-4-activated human chondrocytes: increased expression of lamin A/C and annexins

Activation of Toll-like receptor-4 (TLR-4) in articular chondrocytes increases the catabolic compartment and leads to matrix degradation during the development of osteoarthritis. In this study, we determined the proteomic and genomic alterations in human chondrocytes during lipopolysaccharide (LPS)-induced inflammation to elucidate the underlying mechanisms and consequences of TLR-4 activation. Human chondrocytes were cultured with LPS for 12, 24, and 36 h to induce TLR-4 activation. The TLR-4-induced inflammatory response was confirmed by real-time PCR analysis of increased interleukin-1 beta (IL-1β), interleukin-6 (IL-6), and tumor necrosis factor alpha (TNF-α) expression levels. In TLR-4-activated chondrocytes, proteomic changes were determined by two-dimensional electrophoresis and matrix-assisted laser desorption/ionization-mass spectroscopy analysis, and genomic changes were determined by microarray and gene ontology analyses. Proteomics analysis identified 26 proteins with significantly altered expression levels; these proteins were related to the cytoskeleton and oxidative stress responses. Gene ontology analysis indicated that LPS treatment altered specific functional pathways including ‘chemotaxis’, ‘hematopoietic organ development’, ‘positive regulation of cell proliferation’, and ‘regulation of cytokine biosynthetic process’. Nine of the 26 identified proteins displayed the same increased expression patterns in both proteomics and genomics analyses. Western blot analysis confirmed the LPS-induced increases in expression levels of lamin A/C and annexins 4/5/6. In conclusion, this study identified the time-dependent genomic, proteomic, and functional pathway alterations that occur in chondrocytes during LPS-induced TLR-4 activation. These results provide valuable new insights into the underlying mechanisms that control the development and progression of osteoarthritis.

NecroX-5 protects mitochondrial oxidative phosphorylation capacity and preserves PGC1alpha expression levels during hypoxia/reoxygenation injury

Although the antioxidant and cardioprotective effects of NecroX-5 on various in vitro and in vivo models have been demonstrated, the action of this compound on the mitochondrial oxidative phosphorylation system remains unclear. Here we verify the role of NecroX-5 in protecting mitochondrial oxidative phosphorylation capacity during hypoxia-reoxygenation (HR). Necrox-5 treatment (10 microM) and non-treatment were employed on isolated rat hearts during hypoxia/reoxygenation treatment using an ex vivo Langendorff system. Proteomic analysis was performed using liquid chromatography-mass spectrometry (LC-MS) and non-labeling peptide count protein quantification. Real-time PCR, western blot, citrate synthases and mitochondrial complex activity assays were then performed to assess heart function. Treatment with NecroX-5 during hypoxia significantly preserved electron transport chain proteins involved in oxidative phosphorylation and metabolic functions. NecroX-5 also improved mitochondrial complex I, II, and V function. Additionally, markedly higher peroxisome proliferator-activated receptor-gamma coactivator-1alpha (PGC1alpha) expression levels were observed in NecroX-5-treated rat hearts. These novel results provide convincing evidence for the role of NecroX-5 in protecting mitochondrial oxidative phosphorylation capacity and in preserving PGC1alpha during cardiac HR injuries.

Mitochondrial calcium uniporter inhibition attenuates mouse bone marrow-derived mast cell degranulation induced by beta-1,3-glucan

Mast cells are primary mediators of allergic inflammation. Beta-1,3-glucan (BG) protects against infection and shock by activating immune cells. Activation of the BG receptor induces an increase in intracellular Ca2+, which may induce exocytosis. However, little is known about the precise mechanisms underlying BG activation of immune cells and the possible role of mitochondria in this process. The present study examined whether BG induced mast cell degranulation, and evaluated the role of calcium transients during mast cell activation. Our investigation focused on the role of the mitochondrial calcium uniporter (MCU) in BG-induced degranulation. Black mouse (C57) bone marrow-derived mast cells were stimulated with 0.5 microg/ml BG, 100 microg/ml peptidoglycan (PGN), or 10 microM A23187 (calcium ionophore), and dynamic changes in cytosolic and mitochondrial calcium and membrane potential were monitored. BG-induced mast cell degranulation occurred in a time-dependent manner, and was significantly reduced under calcium-free conditions. Ruthenium red, a mitochondrial Ca2+ uniporter blocker, significantly reduced mast cell degranulation induced by BG, PGN, and A23187. These results suggest that the mitochondrial Ca2+ uniporter has an important regulatory role in BG-induced mast cell degranulation.

NecroX-5 exerts anti-inflammatory and anti-fibrotic effects via modulation of the TNFα/Dcn/TGFβ1/Smad2 pathway in hypoxia/reoxygenation-treated rat hearts

Inflammatory and fibrotic responses are accelerated during the reperfusion period, and excessive fibrosis and inflammation contribute to cardiac malfunction. NecroX compounds have been shown to protect the liver and heart from ischemia-reperfusion injury. The aim of this study was to further define the role and mechanism of action of NecroX-5 in regulating infl ammation and fi brosis responses in a model of hypoxia/reoxygenation (HR). We utilized HR-treated rat hearts and lipopolysaccharide (LPS)-treated H9C2 culture cells in the presence or absence of NecroX-5 (10 µmol/L) treatment as experimental models. Addition of NecroX-5 signifi cantly increased decorin (Dcn) expression levels in HR-treated hearts. In contrast, expression of transforming growth factor beta 1 (TGFβ1) and Smad2 phosphorylation (pSmad2) was strongly attenuated in NecroX-5-treated hearts. In addition, signifi cantly increased production of tumor necrosis factor alpha (TNFα), TGFβ1, and pSmad2, and markedly decreased Dcn expression levels, were observed in LPS-stimulated H9C2 cells. Interestingly, NecroX-5 supplementation effectively attenuated the increased expression levels of TNFα, TGFβ1, and pSmad2, as well as the decreased expression of Dcn. Thus, our data demonstrate potential antiinflammatory and anti-fibrotic effects of NecroX-5 against cardiac HR injuries via modulation of the TNFα/Dcn/TGFβ1/Smad2 pathway.

Mitochondrial pyruvate dehydrogenase phosphatase 1 regulates the early differentiation of cardiomyocytes from mouse embryonic stem cells

Mitochondria are crucial for maintaining the properties of embryonic stem cells (ESCs) and for regulating their subsequent differentiation into diverse cell lineages, including cardiomyocytes. However, mitochondrial regulators that manage the rate of differentiation or cell fate have been rarely identified. This study aimed to determine the potential mitochondrial factor that controls the differentiation of ESCs into cardiac myocytes. We induced cardiomyocyte differentiation from mouse ESCs (mESCs) and performed microarray assays to assess messenger RNA (mRNA) expression changes at differentiation day 8 (D8) compared with undifferentiated mESCs (D0). Among the differentially expressed genes, Pdp1 expression was significantly decreased (27-fold) on D8 compared to D0, which was accompanied by suppressed mitochondrial indices, including ATP levels, membrane potential, ROS and mitochondrial Ca²⁺. Notably, Pdp1 overexpression significantly enhanced the mitochondrial indices and pyruvate dehydrogenase activity and reduced the expression of cardiac differentiation marker mRNA and the cardiac differentiation rate compared to a mock control. In confirmation of this, a knockdown of the Pdp1 gene promoted the expression of cardiac differentiation marker mRNA and the cardiac differentiation rate. In conclusion, our results suggest that mitochondrial PDP1 is a potential regulator that controls cardiac differentiation at an early differentiation stage in ESCs.

Advanced chronic kidney disease: a strong risk factor for Clostridium difficile infection

BACKGROUND/AIMS: It has been suggested that chronic kidney disease (CKD) is a risk factor for Clostridium difficile infection (CDI) and is associated with increased mortality among patients infected with C. difficile. However, recent studies of the clinical impact of CKD on CDI in Asians are still insufficient. We sought to determine the relationship between CKD and CDI in a Korean population. METHODS: This was a single-center, retrospective case-control study. In total, 171 patients with CDI were included as cases and 342 age- and gender-matched patients without CDI were used as controls. We compared the prevalence of CKD in the study sample and identified independent risk factors that could predict the development or prognosis of CDI. RESULTS: Independent risk factors for CDI included stage IV to V CKD not requiring dialysis (odds ratio [OR], 2.90) and end-stage renal disease requiring dialysis (OR, 3.34). Patients with more advanced CKD (estimated glomerular filtration rate < 30) and CDI showed higher in-hospital mortality and poorer responses to the initial metronidazole therapy. CONCLUSIONS: More advanced CKD is an independent risk factor for CDI and is associated with higher in-hospital mortality and poor treatment responses in CDI patients. Thus, in CKD patients, careful attention should be paid to the occurrence of CDI and its management to improve the outcome of CDI.

Intra-abdominal hypertension does not predict renal recovery or in-hospital mortality in critically ill patients with acute kidney injury

BACKGROUND: Although emerging evidence suggests that intra-abdominal hypertension (IAH) is a predictor of the development of acute kidney injury (AKI), it remains unclear whether the presence of IAH is a predictor of prognosis in patients with AKI. The purpose of this study was to assess whether the presence of IAH could predict prognosis in critically ill patients with AKI. The prognostic value of urinary biomarkers was also determined. METHODS: In this prospective observational study, we enrolled 57 patients with established AKI, who were admitted to the intensive care unit between February 2012 and June 2014. IAH was defined as a sustained elevation in intra-abdominal pressure of > or =12 mmHg, in three consecutive measurements performed daily on the first 3 days. Urinary neutrophil gelatinase-associated lipocalin (NGAL), liver-type fatty acid-binding protein, and simplified acute physiology score II score at the time of admission were also examined. RESULTS: IAH was observed in 78.9% of patients. The in-hospital mortality was 21.1%, and renal recovery during hospitalization was achieved in 40.4% of patients. Although high urinary NGAL [odds ratio (OR), 1.015] and liver-type fatty acid-binding protein (OR, 1.003) were found to be independent predictors of renal recovery, IAH was not. High urinary NGAL (OR, 1.003) and a high simplified acute physiology score II score (OR, 1.102) were independent predictors of in-hospital mortality, while IAH or urinary liver-type fatty acid-binding protein was not. CONCLUSION: Although IAH is prevalent in critically ill patients with AKI, it did not predict AKI prognosis. However, urinary NGAL was found to be a useful predictor of both renal recovery and in-hospital mortality.

Renal Klotho expression in patients with acute kidney injury is associated with the severity of the injury

BACKGROUND/AIMS: The potential physiologic roles of Klotho in acute kidney injury (AKI) have recently been demonstrated in animal models. However, to date, there have been no human studies investigating the expression of renal Klotho in AKI. METHODS: We retrospectively collected biopsy specimens and clinical data of AKI patients between January 2001 and December 2012. Klotho expression was determined by immunohistochemical staining, and the clinical-pathological correlation was examined. RESULTS: Among the 34 patients diagnosed with acute tubular necrosis or acute tubulointerstitial nephritis, 21 patients without chronic histological lesions were included. The mean age was 37.3 +/- 18.5 years and the mean peak creatinine level was 8.2 +/- 5.5 mg/dL. In total, 10 patients (47.6%) received temporary renal replacement therapy (RRT); however, 17 patients (81%) showed functional recovery with creatinine levels of < 1.3 mg/dL after 1 month. The intensity of Klotho expression was scored as a percentage of Klotho-positive area. The renal Klotho score showed a significant negative correlation with the initial or peak creatinine level. When the patients were divided into three groups according to the Klotho score (low, middle, high), the low group had a significantly higher peak creatinine level and a more frequent requirement for RRT. However, the Klotho score was not a significant predictor of renal recovery. CONCLUSIONS: The results demonstrated that renal Klotho expression in humans decreased significantly according to the severity of AKI, regardless of the etiology, and that low expression was associated with a poor short-term outcome.

Etiology and outcomes of anuria in acute kidney injury: a single center study

BACKGROUND: It was previously known that anuric acute kidney injury (AKI) is uncommon and its occurrence suggests complete ureteral obstruction, shock, or a major vascular event. As the epidemiology of AKI has significantly changed over the past decade, it is possible that the incidence, etiology, or clinical characteristics of anuric AKI have also changed. METHODS: A prospective cohort study was conducted that included all patients undergoing renal replacement therapy (RRT) for AKI during a 2-year period in a tertiary hospital. Patients were classified as having anuric, oliguric, or nonoliguric AKI based on their volume of urine when RRT started using the modified Acute Kidney Injury Network criteria. RESULTS: Of the 203 patients included in the study, 21.2% met the criteria for anuric AKI. Septic and postoperative AKI were the main causes of anuric AKI, with 60.5% of incidences occurring in hospital. Anuric AKI was associated with a younger age, a lower prevalence of pre-morbid chronic kidney disease and diabetes, more frequent continuous RRT requirement, and multi-organ dysfunction. In addition, patients with anuric AKI had a higher rate of in-hospital mortality and long-term dependence on RRT than patients with nonanuric AKI. CONCLUSION: Anuric AKI is common, with sepsis as the main etiological insult, and is associated with adverse outcomes among patients with AKI who require RRT.

Relationship between pulmonary hypertension, peripheral vascular calcification, and major cardiovascular events in dialysis patients

BACKGROUND: Pulmonary hypertension (PHT) is a recently recognized complication of chronic kidney disease. In this study, we investigated the association between PHT, peripheral vascular calcifications (VCs), and major cardiovascular events. METHODS: In this retrospective study, we included 172 end-stage renal disease (ESRD) patients undergoing dialysis [hemodialysis (HD)=84, peritoneal dialysis=88]. PHT was defined as an estimated pulmonary artery systolic pressure >37 mmHg using echocardiography. The Simple Vascular Calcification Score (SVCS) was measured using plain radiographic films of the hands and pelvis. RESULTS: The prevalence of PHT was significantly higher in HD patients (51.2% vs. 22.7%). Dialysis patients with PHT had a significantly higher prevalence of severe VCs (SVCS> or =3). In multivariate analysis, the presence of severe VCs [odds ratio (OR), 2.68], mitral valve disease (OR, 7.79), HD (OR, 3.35), and larger left atrial diameter (OR, 11.39) were independent risk factors for PHT. In addition to the presence of anemia, severe VCs, or older age, the presence of PHT was an independent predictor of major cardiovascular events in ESRD patients. CONCLUSION: The prevalence of PHT was higher in HD patients and was associated with higher rates of major cardiovascular events. Severe VCs are thought to be an independent risk factor for predicting PHT in ESRD patients. Therefore, in dialysis patients with PHT, careful attention should be paid to the presence of VCs and the occurrence of major cardiovascular events.

The Critical Roles of Zinc: Beyond Impact on Myocardial Signaling

Zinc has been considered as a vital constituent of proteins, including enzymes. Mobile reactive zinc (Zn2+) is the key form of zinc involved in signal transductions, which are mainly driven by its binding to proteins or the release of zinc from proteins, possibly via a redox switch. There has been growing evidence of zinc's critical role in cell signaling, due to its flexible coordination geometry and rapid shifts in protein conformation to perform biological reactions. The importance and complexity of Zn2+ activity has been presumed to parallel the degree of calcium's participation in cellular processes. Whole body and cellular Zn2+ levels are largely regulated by metallothioneins (MTs), Zn2+ importers (ZIPs), and Zn2+ transporters (ZnTs). Numerous proteins involved in signaling pathways, mitochondrial metabolism, and ion channels that play a pivotal role in controlling cardiac contractility are common targets of Zn2+. However, these regulatory actions of Zn2+ are not limited to the function of the heart, but also extend to numerous other organ systems, such as the central nervous system, immune system, cardiovascular tissue, and secretory glands, such as the pancreas, prostate, and mammary glands. In this review, the regulation of cellular Zn2+ levels, Zn2+-mediated signal transduction, impacts of Zn2+ on ion channels and mitochondrial metabolism, and finally, the implications of Zn2+ in health and disease development were outlined to help widen the current understanding of the versatile and complex roles of Zn2+.

Polymicrobial Peritonitis with Lactococcus lactis in a Peritoneal Dialysis Patient / 전남의대학술지

Lactococcus lactis (L. lactis) is an important gram-positive bacterium in dairy products. It is a rare cause of opportunistic infections with only four cases of Lactococcus peritoneal dialysis (PD) peritonitis reported in the literature. In Korea, L. lactis infection was first reported in a liver abscess patient in 2010; however, PD peritonitis with Lactococcus has not been reported in Korea. Recently, we experienced a case of Lactococcus-associated polymicrobial PD peritonitis. The patient was initially managed with broad-coverage antibiotics; however, owing to a poor response, the PD catheter was removed and the patient was switched to hemodialysis. We discuss this case and review the literature.

Kinesin Spindle Protein Inhibition in Translational Research

The kinesin superfamily is a class of motor proteins moving along microtubule filaments and playing essential roles in mitosis of eukaryotic cells. In the cancer biology, mitotic activity is an essential factor for development and metastasis of various cancers. Therefore, the inhibition of kinesin activity is suggested as an alternative cancer therapy. Accumulated clinical evidences have proved the potency of kinesin inhibitors in cancer treatments. In this review, we provided an overview of kinesins that play a critical role in the pathophysiology of various cancers and described the beneficial vs. side effects of their inhibitors that have been tested in both basic science and clinical studies.

Polymicrobial Peritonitis with Lactococcus lactis in a Peritoneal Dialysis Patient / 전남의대학술지

Lactococcus lactis (L. lactis) is an important gram-positive bacterium in dairy products. It is a rare cause of opportunistic infections with only four cases of Lactococcus peritoneal dialysis (PD) peritonitis reported in the literature. In Korea, L. lactis infection was first reported in a liver abscess patient in 2010; however, PD peritonitis with Lactococcus has not been reported in Korea. Recently, we experienced a case of Lactococcus-associated polymicrobial PD peritonitis. The patient was initially managed with broad-coverage antibiotics; however, owing to a poor response, the PD catheter was removed and the patient was switched to hemodialysis. We discuss this case and review the literature.

Combination treatment with 2-methoxyestradiol overcomes bortezomib resistance of multiple myeloma cells

Bortezomib is a proteasome inhibitor used for the treatment of relapsed/refractory multiple myeloma (MM). However, intrinsic and acquired resistance to bortezomib has already been observed in MM patients. In a previous report, we demonstrated that changes in the expression of mitochondrial genes lead to changes in mitochondrial activity and bortezomib susceptibility or resistance, and their combined effects contribute to the differential sensitivity or resistance of MM cells to bortezomib. Here we report that the combination treatment of bortezomib and 2-methoxyestradiol (2ME), a natural estrogen metabolite, induces mitochondria-mediated apoptotic cell death of bortezomib-resistant MM KMS20 cells via mitochondrial reactive oxygen species (ROS) overproduction. Bortezomib plus 2ME treatment induces a higher level of cell death compared with treatment with bortezomib alone and increases mitochondrial ROS and Ca2+ levels in KMS20 cells. Pretreatment with the antioxidant N-acetyl-L-cysteine scavenges mitochondrial ROS and decreases cell death after treatment with bortezomib plus 2ME in KMS20 cells. Moreover, we observed that treatment with bortezomib plus 2ME maintains the activation of c-Jun N-terminal kinase (JNK) and mitogen-activated protein kinase kinase kinase 4/7 (MKK4/7). Collectively, combination treatment with bortezomib and 2ME induces cell death via JNK-MKK4/7 activation by overproduction of mitochondrial ROS. Therefore, combination therapy with specific mitochondrial-targeting drugs may prove useful to the development of novel strategies for the treatment of bortezomib-resistant MM patients.

Proteomic Changes in Rat Gastrocnemius Muscle After Botulinum Toxin A Injection

OBJECTIVE: To observe the changes in protein expression induced by botulinum toxin A (BoNT-A) injection and to characterize the molecular and cellular action of mechanisms of BoNT-A injection on skeletal muscles using proteomic elements as biomarkers. METHODS: BoNT-A was injected into left gastrocnemius muscles of 12 Sprague-Dawley rats (2 months of age) at a dosage of 5 units/kg body weight. For the controls same volume of normal saline was injected to right gastrocnemius muscle of each rat. Muscle samples were obtained at 4 time points (3 rats per time point): 3, 7, 14, and 56 day post-injection. To reveal the alterations in muscle protein, we performed 2-dimensional electrophoresis (2DE) and compared Botox group and normal saline group at each time point. Altered protein spots in 2DE were identified using matrix-assisted laser desorption/ionization-time-of-flight mass spectrometer (MALDI-TOF MS) proteomics analysis. RESULTS: Compared with normal saline group, 46 protein spots showed changed protein expression. Twelve protein spots demonstrated increased volume and 34 protein spots demonstrated decreased volume. Among spots of decreased volume, 17 spots showed statistically significant differences. Thirty-eight identified proteins were associated with alterations in energy metabolism, muscle contractile function, transcription, translation, cell proliferation, and cellular stress response. CONCLUSION: BoNT-A gives influences on muscle contractile function and energy metabolism directly or indirectly besides neurotoxic effects. Proteomic expression provides better understanding about the effect of BoNT-A on skeletal muscle.