scholarly journals Cyclophosphamide treatment evoked side effect on skeletal muscle actin, monitored by DSC

2021 ◽  
Author(s):  
Péter Farkas ◽  
Dávid Szatmári ◽  
Franciska Könczöl ◽  
Dénes Lőrinczy
Keyword(s):  
Skeletal Muscle ◽  
Side Effect ◽  
Main Peak ◽  
Muscle Actin ◽  
Scanning Calorimetry ◽  
Drug Induced ◽  
Cyclophosphamide Treatment ◽  
Intact Muscle ◽  
Binding Cleft

AbstractSeveral kind of drugs—used in cancer treatments—such as cyclophosphamide (CP) can also trigger a disease classified as toxic polyneuropathy. Polyneuropathy is a simultaneous malfunction of several peripheral nerves, typical side effect of a cancer therapy. In our previous study, we used CP treated in vitro animal model (Guinea pig) with a comparable dosage and time handling of human protocol to show evidences of this drug-induced effects. We could show a dose-dependent difference between in Tm and ΔHcal of untreated and treated samples assigned to their intact muscle and nerve, blood plasma and red blood cells. In our current study we analyze this side effect on skeletal muscle actin (prepared from m. psoas of rabbit) by DSC (differential scanning calorimetry), to follow the possible consequence of drug treatment on the “activator” of muscle contraction. We have demonstrated that run of DSC curves, Tms together with the ΔHcal exhibit clear CP effect. In case of Ca2+ G actin it is manifested in a well separated second high denaturing temperature as a consequence of CP binding into the cleft. This way the nucleotide binding cleft with subdomains 1 and 3 becomes less flexible, indicating clear sensitivity to CP treatment. In F-actin samples, the main peak represents the thermal denaturation of subdomains 1 and 3, and the increased calorimetric enthalpy administrating Ca2+ as well as CP refers to a more rigid structure. These alterations can be the molecular background in the malfunction of muscle in case of polyneuropathy after CP treatment.

scholarly journals Cyclophosphamide treatment evoked side effects on skeletal muscle monitored by DSC

2020 ◽  
Vol 142 (5) ◽  
pp. 1897-1901
Author(s):  
Dénes Lőrinczy
Keyword(s):  
Skeletal Muscle ◽  
Locomotor Activity ◽  
Side Effects ◽  
Side Effect ◽  
Muscle Protein ◽  
Scanning Calorimetry ◽  
Drug Induced ◽  
Severe Problem ◽  
Cyclophosphamide Treatment

AbstractPolyneuropathy is defined as a simultaneous malfunction of several peripheral nerves, which could be a side effect of cancer therapy as well. Many kinds of drugs, supposedly cyclophosphamide, also can induce a disease classified as toxic polyneuropathy. It is well known that a severe problem in the locomotor activity can join to it. Recently, we have no enough information about the attacked points in the structure of muscle proteins, as well as about the change in the interaction of myosin actin. In the present study, we analyse this side effect on skeletal muscle (m. gastrocnemius) by differential scanning calorimetry (DSC), as an established thermal analysis method, to follow the possible consequence of drug treatment in the most important muscle protein. We used cyclophosphamide-treated in vitro animal model (guinea pig) with a comparable dosage and time handling of human protocol to show evidences of this drug-induced effects. According to our results, we could show a dose-dependent difference between thermal parameters (denaturation temperature and calorimetric enthalpy) of untreated and treated samples assigned to their contractile proteins (actin and myosin), which can be detected by DSC. It proved that we can create new possibilities in the detection and prognosis of expected and unwanted side effects of cyclophosphamide, such as change of locomotor activity joined to polyneuropathy.

scholarly journals Tissue-Engineered Human Myobundle System as a Platform for Evaluation of Skeletal Muscle Injury Biomarkers

2020 ◽  
Vol 176 (1) ◽  
pp. 124-136
Author(s):  
Alastair Khodabukus ◽  
Amulya Kaza ◽  
Jason Wang ◽  
Neel Prabhu ◽  
Richard Goldstein ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Muscle Injury ◽  
Culture Media ◽  
Contractile Function ◽  
Significant Loss ◽  
Drug Induced ◽  
Skeletal Muscle Injury ◽  
Specificity And Sensitivity ◽  
Novel Biomarkers

Abstract Traditional serum biomarkers used to assess skeletal muscle damage, such as activity of creatine kinase (CK), lack tissue specificity and sensitivity, hindering early detection of drug-induced myopathies. Recently, a novel four-factor skeletal muscle injury panel (MIP) of biomarkers consisting of skeletal troponin I (sTnI), CK mass (CKm), fatty-acid-binding protein 3 (Fabp3), and myosin light chain 3, has been shown to have increased tissue specificity and sensitivity in rodent models of skeletal muscle injury. Here, we evaluated if a previously established model of tissue-engineered functional human skeletal muscle (myobundle) can allow detection of the MIP biomarkers after injury or drug-induced myotoxicity in vitro. We found that concentrations of three MIP biomarkers (sTnI, CKm, and Fabp3) in myobundle culture media significantly increased in response to injury by a known snake venom (notexin). Cerivastatin, a known myotoxic statin, but not pravastatin, induced significant loss of myobundle contractile function, myotube atrophy, and increased release of both traditional and novel biomarkers. In contrast, dexamethasone induced significant loss of myobundle contractile function and myotube atrophy, but decreased the release of both traditional and novel biomarkers. Dexamethasone also increased levels of matrix metalloproteinase-2 and -3 in the culture media which correlated with increased remodeling of myobundle extracellular matrix. In conclusion, this proof-of-concept study demonstrates that tissue-engineered human myobundles can provide an in vitro platform to probe patient-specific drug-induced myotoxicity and performance assessment of novel injury biomarkers to guide preclinical and clinical drug development studies.

scholarly journals Extraction Protocols for Individual Zebrafish's Ventricle Myosin and Skeletal Muscle Actin for In vitro Motility Assays

2017 ◽  
Vol 8 ◽  
Author(s):  
Lisa-Mareike Scheid ◽  
Cornelia Weber ◽  
Nasrin Bopp ◽  
Matias Mosqueira ◽  
Rainer H. A. Fink
Keyword(s):  
Skeletal Muscle ◽  
Muscle Actin ◽  
In Vitro Motility

scholarly journals Muscle actin filaments bind pituitary secretory granules in vitro.

1977 ◽  
Vol 73 (1) ◽  
pp. 78-87 ◽  
Author(s):  
R E Ostlund ◽  
J T Leung ◽  
D M Kipnis
Keyword(s):  
Skeletal Muscle ◽  
Metal Ions ◽  
Secretory Granule ◽  
Actin Filaments ◽  
Anterior Pituitary ◽  
Secretory Granules ◽  
Muscle Actin ◽  
Viscous Solutions ◽  
Styrene Butadiene

Hog anterior pituitary secretory granules sediment at 3,000 g. When rat or rabbit skeletal muscle actin filaments are present with the granules, the sedimentation decreases markedly. Depolymerized actin or viscous solutions of Ficoll and collagen have no effect on granule sedimentation. With this assay, actin filaments bind secretory granules (consisting of the proteinaceous core plus limiting membrane), secretory granule membranes, mitochondria, artificial lecithin liposomes, and styrene-butadiene microspheres, but have little or no interaction with membrane-free secretory granule cores and albumin microspheres. A secretory granule-actin complex sedimentable between 3,000 g and 25,000 g can be isolated. Metal ions, nucleotides, salts, dithiothreitol, or pretreatment of the granules with trypsin do not destroy the binding, which appears to be a lipophilic interaction.

Blockade of Chemotherapy-Induced Thrombocytopenia by Bax Inhibiting Peptides (BIPs) in Mouse Model.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 281-281
Author(s):  
Jose A. Gomez ◽  
Vivian Gama ◽  
Tomoyuki Yoshida ◽  
Michelle Stapleton ◽  
Cathy Paddock ◽  
...  
Keyword(s):  
Platelet Count ◽  
Side Effect ◽  
Bleeding Time ◽  
Caspase Inhibitor ◽  
Wild Type ◽  
Drug Induced ◽  
Induced Apoptosis ◽  
Platelet Formation

Abstract Thrombocytopenia is a major side effect of cancer chemotherapy producing bleeding problems as well as cancer treatment delays. To preclude it, the platelets progenitors megakaryocytes need to be protected from drug-induced apoptosis. According to previous reports, anti-apoptotic molecules such as Bcl-2, Bcl-xL, and caspase inhibitors were found to interfere platelet formation by megakaryocytes, thus these apoptosis inhibitors are not appropriate therapeutic candidates for the prevention of drug-induced thrombocytopenia. Recently we developed a new type of cell permeable apoptosis inhibitor, Bax Inhibiting Peptides (BIPs), which are derived from the Bax binding domain of Ku70. Bax is a key mediator of apoptosis, and Ku70 is a multifunctional protein playing roles in cell survival and DNA repair. In the present study, we determined whether BIPs prevent chemotherapy-induced thrombocytopenia both in vitro and in vivo. Importantly, there are several cancer cell types that acquired mutations in genes of Bax and/or Bax activating factors. Therefore, we hypothesize that protection of non-tumorigenic essential cells (such as megakaryocytes) by BIPs is beneficial for patients suffering from Bax-negative cancer. For in vitro study, Dami cells (human pro-megakaryocyte cell line) and mouse primary cultured megakaryocytes were used. BIPs significantly inhibited apoptosis induction by anti-cancer drugs such as etoposide, cisplatin, doxorubicin, or paclitaxel. Importantly, BIPs did not interfere the activity of megakaryocytes to produce platelet-like particle in cell culture, suggesting that BIPs protect megakaryocytes from drug-induced apoptosis without suppressing their platelets production. For in vivo study, C57BL/6J mice and Bax-/- mice (backcrossed with C57BL/6J) were used. Etoposide (150 mg/kg, i.p.) induced a significant decrease of platelet count in whole blood within 2 days after the drug administration in wild type C57BL/6J mice. Pretreatment of mice with BIPs (166 mg/kg, i.p., 30 min before etoposide injection) blocked etoposide-induced platelet count decrease in these wild type mice. The same dose of etoposide did not induce significant platelet count decrease in Bax-/- mice. These results suggest that chemotherapy-induced thrombocytopenia, at least in part, is due to the stimulation of Bax-mediated cell death probably both in megakaryocytes and platelets. In our preliminary experiments, three times injection of caspase inhibitor (z-VAD-fmk, 166 mg/kg, i.p. every 24 hrs) caused prolongation of bleeding time from the wound (tail cut). Thus, as previously reported by other groups using in vitro system, caspase inhibitor may have its own side effect to inhibit platelet formation or/and activation in vivo. On the other hand, BIPs (166 mg/kg, 3 times every 24 hrs, i.p.) did not extend bleeding time from the tail. Taken together, BIP may become a useful supplemental therapeutic agent to block chemotherapy-induced thrombocytopenia.

Ultrastructural autoradiography of L6 skeletal-muscle cells exposed to a tritiated macrolide antibiotic (LY237216) in vitro

1992 ◽  
Vol 50 (1) ◽  
pp. 612-613
Author(s):  
S.L. White ◽  
C.B. Jensen ◽  
D.D. Giera ◽  
D.A. Laska ◽  
M.N. Novilla ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Quantitative Analysis ◽  
Macrolide Antibiotic ◽  
Circle Method ◽  
Apical Surface ◽  
Polycarbonate Membrane ◽  
L6 Cells ◽  
Low Viscosity ◽  
Erythromycin A

In vitro exposure to LY237216 (9-Deoxo-11-deoxy-9,11-{imino[2-(2-methoxyethoxy)ethylidene]-oxy}-(9S)-erythromycin), a macrolide antibiotic, was found to induce cytoplasmic vacuolation in L6 skeletal muscle myoblast cultures (White, S.L., unpubl). The present study was done to determine, by autoradiographic quantitative analysis, the subcellular distribution of 3H-LY237216 in L6 cells.L6 cells (ATCC, CRL 1458) were cultured to confluency on polycarbonate membrane filters (Millipore Corp., Bedford, MA) in M-199 medium (GIBCO® Labs) with 10% fetal bovine serum. The cells were exposed from the apical surface for 1-hour to unlabelled-compound (0 μCi/ml) or 50 (μCi/ml of 3H-LY237216 at a compound concentration of 0.25 mg/ml. Following a rapid rinse in compound-free growth medium, the cells were slam-frozen against a liquid nitrogen cooled, polished copper block in a CF-100 cryofixation unit (LifeCell Corp., The Woodlands, TX). Specimens were dried in the MDD-C Molecular Distillation Drier (LifeCell Corp.), vapor osmicated and embedded in Spurrs low viscosity resin. Ultrathin sections collected on formvar coated stainless steel grids were counter-stained, then individually mounted on corks. A monolayer of Ilford L4 nuclear emulsion (Polysciences, Inc., Warrington, PA) was placed on the sections, utilizing a modified “loop method”. The emulsions were exposed for 7-weeks in a light-tight box at 4°C. Autoradiographs were developed in Microdol-X developer and examined on a Philips EM410LS transmission electron microscope. Quantitative analysis of compound localization employed the point and circle approach of Williams; incorporating the probability circle method of Salpeter and McHenry.

A Hexagonal (II) Phase Phospholipid Neutralization Assay for Lupus Anticoagulant Identification

1993 ◽  
Vol 70 (05) ◽  
pp. 787-793 ◽  
Author(s):  
Douglas A Triplett ◽  
Linda K Barna ◽  
Gail A Unger
Keyword(s):  
Hemophilia A ◽  
Clinical Laboratory ◽  
Neutralization Assay ◽  
Confirmatory Test ◽  
Specific Factor ◽  
Clinical Settings ◽  
Drug Induced ◽  
Variable Screening ◽  
Routine Clinical Laboratory

SummaryLupus anticoagulants (LAs) are immunoglobulins (IgG, IgM, or both) which interfere with in vitro phospholipid (PL) dependent tests of coagulation (e.g. APTT, dilute PT, dilute Russell Viper Venom Time). These antibodies may be identified in a wide variety of clinical settings. With the exception of heparinized patient samples, the presence of LAs is often the most common cause of an unexplained APTT in a routine clinical laboratory. The diagnosis of LAs is difficult due to variable screening reagent sensitivity and intrinsic heterogeneity of LAs. Recently, Rauch and colleagues have shown human monoclonal hybridoma LAs were inhibited by hexagonal (II) phase PLs. In contrast, lamellar phase PLs had no effect. We have evaluated a new assay system, Staclot LA®, which utilizes a hexagonal (II) phase PL (egg phosphatidylethanolamine [EPE]) as a confirmatory test for LAs. Plasma samples from the following patient populations were studied: LA positive, heparinized, oral anticoagulated, hemophilia A and B, and specific factor inhibitors (factors V, VIII, IX). Unlike previous studies, the LA positive patients were a mixed population including: autoimmune diseases, drug-induced, and post-infection. Our findings confirm the specificity of hexagonal (II) phase PL neutralization of LAs.

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