Journal of the Neurological Sciences 345 (2014) 239–243
Contents lists available at ScienceDirect
Journal of the Neurological Sciences journal homepage: www.elsevier.com/locate/jns
Short communication
Polymyositis in solid organ transplant recipients receiving tacrolimus Gaetano Vattemi a, Matteo Marini a, Marzia Di Chio b, Maria Colpani c, Valeria Guglielmi a, Giuliano Tomelleri a,⁎ a b c
Department of Neurological and Movement Sciences, Section of Clinical Neurology, University of Verona, Italy Department of Medicine and Public Health, Section of Pharmacology, University of Verona, Verona, Italy Department of Gastroenterology, Liver Transplantation Unit, “Ospedali Riuniti”, Bergamo, Italy
a r t i c l e
i n f o
Article history: Received 4 January 2014 Received in revised form 17 June 2014 Accepted 15 July 2014 Available online 6 August 2014 Keywords: Tacrolimus Solid organ transplantation Muscle toxicity Polymyositis Calcineurin inhibitor T cells
a b s t r a c t Tacrolimus, also known as FK506, is an immunosuppressive agent widely used for the prevention of acute allograft rejection in organ transplantation and for the treatment of immunological diseases. This study reports two male patients who underwent solid organ transplantation (liver and kidney). After transplant, the patients received continuous immunosuppressive therapy with oral tacrolimus and later presented clinical manifestations and laboratory signs of myopathy. Muscle biopsies of both patients clearly documented an inflammatory myopathy with the histological features of polymyositis including CD8+ T cells which invaded healthy muscle fibers and expressed granzyme B and perforin, many CD68+ macrophages and MHC class I antigen upregulation on the surface of most fibers. Because of the temporal association while receiving tacrolimus and since other possible causes for myopathy were excluded, the most likely cause of polymyositis in our patients was tacrolimus toxicity. We suggest that patients on tacrolimus should be carefully monitored for serum CK levels and clinical signs of muscle disease. © 2014 Elsevier B.V. All rights reserved.
1. Introduction
2. Case reports
Tacrolimus (FK506) is a calcineurin inhibitor widely used as potent immunosuppressant agent for the prevention of rejection in solid organ transplantation [1,2]. The drug was also introduced in the therapy of several autoimmune diseases including myasthenia gravis and difficult-to-treat cases of polymyositis [3]. In addition, calcineurin inhibitors have been proposed among second-line agents for treatment of myositis and polymyositis, the only “distinctive” neurological manifestations in chronic graft-versus-host diseases (GVHD), a syndrome characterized by immune-mediated multisystemic inflammation occurring after allogeneic hematopoietic stem cell transplantation (HSCT) [4,5]. One study reported a renal transplant patient who had, 10 months after he was started on tacrolimus, an inflammatory myopathy which resolved by its withdrawal [6]. We describe a patient who developed myositis after immunosuppressive treatment with FK506, and performed a detailed analysis of the pathological features of muscle biopsy from this patient and the previous one.
2.1. Patient 1
⁎ Corresponding author at: Department of Neurological and Movement Sciences, Section of Clinical Neurology, University of Verona, P.le L.A. Scuro 10, 37134 Verona, Italy. Tel.: +39 045 8074285; fax: +39.045.8027492. E-mail address: [email protected] (G. Tomelleri).
http://dx.doi.org/10.1016/j.jns.2014.07.036 0022-510X/© 2014 Elsevier B.V. All rights reserved.
This 62-year-old male patient underwent living-donor liver transplantation for alcohol-related cirrhosis two years before. After liver transplant, the patient received continuous immunosuppressive therapy with 1 mg of oral tacrolimus per day. His past medical history disclosed hypertension and diabetes and therapy included low dose of insulin, furosemide, allopurinol and ursodeoxycholic acid. Twenty-one months after the transplantation, the patient developed bile duct stenosis and investigation showed increased levels of serum creatine kinase (CK) (1880 U/L; normal b 200), lactate dehydrogenase (852 U/L; normal b 450), aspartate aminotransferase (160 U/L; normal b 50) and alanine aminotransferase (171 U/L; normal b 50). CK levels were persistently elevated (1600 U/L); electroneurography and needle electromyography documented an axonal peripheral neuropathy. At the time of our observation, three months later, on clinical examination the patient squatted and rose from the floor with difficulty and weakness of the thigh flexor (MRC 4) and quadriceps (MRC 4) muscles was present. Deep tendon reflexes were absent in the arms and legs and vibration sense was lost in the legs. At that time, an open muscle biopsy was obtained from the vastus lateralis. Two months later, the patient underwent surgery for bile duct stenosis but then he developed invasive aspergillosis and two weeks later he died.
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G. Vattemi et al. / Journal of the Neurological Sciences 345 (2014) 239–243
2.2. Patient 2
3.2. Histology and histochemistry
Clinical data have been previously reported [6]. Briefly a 53-year-old man with end-stage renal disease caused by chronic glomerulonephritis underwent a renal transplantation at age 40 and received a second renal graft six years later. A combination treatment of cyclosporine A and azathioprine was started after the second transplant. At age 52 the patient's immunosuppression was switched to tacrolimus, initially 5 mg per day, and within a month 4 mg daily because of high drug serum level. Ten months later the patient complained of muscle pain and laboratory investigations documented a mild increase of CK value, about two–three fold normal value. Because of persistent CK elevations over seven months, an open biopsy of the vastus lateralis muscle was performed. Tacrolimus was then withdrawn followed by resolution of the patient's symptoms and normalization of CK level. The patient was hepatitis C virus (HCV) positive with a viral load of 12.79 MEq/ml two months before muscle biopsy and he was treated with ribavirin which was stopped one month before the elevation of CK.
Serial 8-μm-thick cryosections were stained with standard histological and histochemical methods including hematoxylin and eosin, modified Gomori trichrome, ATPase (pH 4.3, 4.6 and 10.4), succinate dehydrogenase (SDH), cytochrome c oxidase (COX), nicotinamide adenine dinucleotide-tetrazolium reductase (NADH-TR), periodic acid Schiff, Sudan black stains and acid phosphatase.
3. Materials and methods 3.1. Muscle biopsies Muscle biopsies were obtained for diagnostic purposes with written informed consent and the study was approved by the local ethical board.
3.3. Immunohistochemistry and confocal immunofluorescence microscopy Immunohistochemical studies were performed on serial 6.5-μm-thick transverse muscle sections using the following wellcharacterized anti-human antibodies: anti-CD3, anti-CD8, anti-CD20, anti-CD25, anti-CD30, anti-CD57, anti-CD68, anti-MHC (major histocompatibility complex) class I, anti-Granzyme B and anti-Perforin. The reactions were revealed by peroxidase or immunofluorescence methods. Double immunofluorescence was performed using antibodies to CD3 and CD68 in combination with an antibody to CD8. Confocal images were acquired with Zeiss LSM 510 confocal microscope. To control staining specificity the primary antibody was omitted or replaced with nonimmune sera at the same concentration.
Fig. 1. Light microscopy of the muscle biopsies. Histological findings of patient 1 (A–D) and of patient 2 (E and F). A–C (hematoxylin and eosin): Two small endomysial inflammatory infiltrates surrounding and invading healthy muscle fibres (A and B); two necrotic muscle fibers, one of which is entirely invaded by inflammatory cells (B and C); a small basophilic regenerating fiber in C. D: Cytochrome c oxidase shows several COX negative muscle fibres. E, F (hematoxylin and eosin): Three endomysial inflammatory infiltrates surrounding non-necrotic muscle fibers.
G. Vattemi et al. / Journal of the Neurological Sciences 345 (2014) 239–243
4. Results Muscle biopsy from patient 1 documented scattered necrotic fibers invaded by mononuclear cells, basophilic regenerating fibers and small endomysial inflammatory infiltrates invading healthy muscle fibers (Fig. 1 A–C). ATPase reactions showed normal differentiation and distribution of muscle fibers; in addition, sparse lobulated fibers and several COX negative fibers were present by oxidative reactions (Fig. 1D). Inflammatory cells were predominantly composed of CD3 + CD8 + T cells (82% of CD3 + T cells were CD8 +) and CD68 + macrophages (Fig. 2, panels A and B); only single CD20+ B cells were present and CD57+ NK cells were absent. In addition, a few CD25+ T cells and rare CD30+ cells were detected. The cytotoxic CD8+ T cells invading nonnecrotic muscle fibers expressed granzyme B and perforin (Fig. 3A). Muscle fibers invaded by CD8+ T cells showed a sarcolemmal expression of MHC class I antigen, which could be found on several more muscle fibers (Fig. 3B).
241
On muscle biopsy of patient 2 small mononuclear inflammatory cells at endomysial sites were observed; these endomysial inflammatory cells focally surrounded and occasionally invaded non-necrotic muscle fibers (Fig. 1 E,F). The inflammatory infiltrates mainly consisted of CD3 + CD8 + T cells (90% of CD3 + T cells were CD8 +) and CD68 + macrophages (Fig. 2 panel C); neither CD20 + B cells nor CD57 + NK cells were identified. Single CD25 + T cells and CD30 + cells were found. Furthermore, MHC class I antigen was upregulated on the sarcolemma of the majority of the muscle fibers, and granzyme B and perforin were expressed (Fig. 3 C,D). 5. Discussion Tacrolimus is a commonly used T cell-targeted immunosuppressant [1,2]. Adverse effects associated with tacrolimus are mainly dosedependent and include nephrotoxicity, neurotoxicity, hypertension, rhabdomyolysis, altered glucose metabolism, gastrointestinal
Fig. 2. Confocal fluorescence microscopical images. Panel A (patient 1): CD3+ T cells (green) surrounding two non-necrotic muscle fibers are mainly CD8+ T cells (red). Panel B (patient 1): a necrotic muscle fiber invaded by CD68+ macrophages (green) and a healthy fiber surrounded by CD8+ T cells (red). Panel C (patient 2): CD3+ (green) and CD8+ (red) T cells surrounding a non-necrotic muscle fiber.
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G. Vattemi et al. / Journal of the Neurological Sciences 345 (2014) 239–243
Fig. 3. Immunohistochemistry of the muscle biopsies. A, C: Granzyme B staining in muscle biopsy of both patients (A, patient 1 and C, patient 2). B, D: MHC class I antigen is expressed on the sarcolemma of several muscle fibers in both patients (B, patient 1 and D, patient 2).
disturbances, increased susceptibility to infection and malignancy [7]. A recent paper raised the possibility that the drug could induce muscle toxicity [6]. We have report herein two transplant patients who complained of myalgia or muscle weakness, previously described among the side effects of tacrolimus, and had a histologically proven inflammatory myopathy while on immunosuppressive therapy with tacrolimus. The inflammatory myopathy has the features similar to those seen in polymyositis, including CD8 + T cells which invade non-necrotic muscle fibers that express MHC class I antigen [8]. The autoinvasive endomysial CD8+ T cells contain granzyme B and perforin, and MHC class I antigen is ubiquitously upregulated on the surface of most fibers. In view of the temporal association while receiving tacrolimus and since no other cause for myopathy could be determined, we believe that polymyositis in our patients was most likely due to toxicity from tacrolimus. Patient 1 also had clinical and laboratory evidence suggestive of an axonal neuropathy. In our patient diabetes mellitus and chronic alcohol intake were recognized predisposing factors for peripheral neuropathy. In addition, FK506 has been reported to be associated with neuropathy [9]. Although tacrolimus-related neuropathy mainly resembles chronic inflammatory demyelinating polyradiculoneuropathy, patients who developed axonal neuropathy have also been reported [10]. Patient 2 also had HCV infection that in the literature has been reported to be associated with myositis [11]. However, it is unlikely that in our patient the muscle disease was virus-related since the withdrawal of tacrolimus led to the serum CK normalization and the disappearance of muscle symptoms. Skeletal muscle is highly exposed to circulating drugs due to its large global mass and its high metabolic rate and blood flow [12]. Therefore several drugs have been identified as causing toxicity to the muscle tissue [12,13]. Drug-induced inflammatory myopathy has only rarely been
described and offending agents include statins, fibrate, hydroxyurea, penicillamine, omeprazole, niflumic acid, phenytoin, tegafur, interferon alfa, silicon gel, bacilli Calmet-Guerin (BCG) tuberculosis vaccine, alfusozin, the antifungal agent terbinafine [12,13]. At the time of onset of inflammatory myopathy, our patients were not receiving any of all the above drugs. Patient 1 also had histological signs of mitochondrial myopathy suggested by several muscle fibers with COX deficiency. The patient was not treated with any drug inducing mitochondrial toxicity including nucleoside analog reverse transcriptase inhibitors, statins and amiodarone [12,13]. Interestingly, data from previous studies support an adverse effect of tacrolimus on mitochondrial respiratory function [14,15]. On the other hand mitochondrial abnormalities can occur in idiopathic inflammatory myopathies including polymyositis [16]. From the above consideration we reasoned that tacrolimus significantly contributed to the occurrence of inflammatory myopathy in our patients. Paradoxically, an immunosuppressive agent that inhibits T cell activation and proposed for the treatment of refractory polymyositis seems to be related to, or possibly induce T cell-mediated myositis. The mechanism by which tacrolimus might determine deleterious effect on muscle tissue is unknown, but could be comparable to those suggested for other drugs causing an inflammatory myopathy [13]. In addition, tacrolimus prevents programmed cell death of activated T cells, thereby disturbing T cell subset balance and causing an abnormally high number of autoreactive T cells [17]. Moreover, in an in vitro model of T cell activation tacrolimus had only a moderate effect on granzyme B production by CD8+ T cells [18]. We conclude that patients receiving tacrolimus should be made aware of the early muscular symptoms and carefully monitored for CK levels in order to consider reduced dosages of the drug or alternative immunosuppressive therapy.
G. Vattemi et al. / Journal of the Neurological Sciences 345 (2014) 239–243
Funding and conflict of interest statement The authors declare that they have no conflicts of interest, and no funding was required for this work. References [1] Tocci MJ, Matkovich DA, Collier KA, Kwok P, Dumont F, Lin S, et al. The immunosuppressant FK506 selectively inhibits expression of early T cell activation genes. J Immunol 1989;143:718–26. [2] Jang HJ, Kim SC, Han DJ. Tacrolimus for rescue therapy in refractory renal allograft rejection. Transplant Proc 2000;32:1765–6. [3] Schneider-Gold C, Hartung HP, Gold R. Mycophenolate mofetil and tacrolimus: new therapeutic options in neuroimmunological diseases. Muscle Nerve 2006;34:284–91. [4] Wolff D, Schleuning M, von Harsdorf S, Bacher U, Gerbitz A, Stadler M, et al. Consensus conference on clinical practice in chronic GVHD: second-line treatment of chronic graft-versus-host disease. Biol Blood Marrow Transplant 2011;17:1–17. [5] Filipovich AH, Weisdorf D, Pavletic S, Socie G, Wingard JR, Lee SJ, et al. National Institutes of Health consensus development project on criteria for clinical trials in chronic graft-versus-host disease: I. Diagnosis and staging working group report, Biol Blood Marrow Transplant 2005;11:945–56. [6] Orlandi V, Campieri C, Mosconi G, D'Arcangelo GL, Feliciangeli G, Scolari MP, et al. Tacrolimus-associated myositis: a case report in a renal transplant patient. Transplant Proc 2004;36:708–10. [7] Plosker GL, Foster RH. Tacrolimus: a further update of its pharmacology and therapeutic use in the management of organ transplantation. Drugs 2000;59: 323–89.
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[8] Dalakas MC. Review: an update on inflammatory and autoimmune myopathies. Neuropathol Appl Neurobiol 2011;37:226–42. [9] Umapathi T, Chaudhry V. Toxic neuropathy. Curr Opin Neurol 2005;18:574–80. [10] De Weerdt A, Claeys KG, De Jonghe P, Ysebaert D, Chapelle T, Roeyen G, et al. Tacrolimus-related polyneuropathy: case report and review of the literature. Clin Neurol Neurosurg 2008;110:291–4. [11] Ito H, Nagano M, Nakano S, Shigeyoshi Y, Kusaka H. In situ identification of hepatitis C virus RNA in muscle. Neurology 2005;64:1073–5. [12] Klopstock T. Drug-induced myopathies. Curr Opin Neurol 2008;21:590–5. [13] Dalakas MC. Toxic and drug-induced myopathies. J Neurol Neurosurg Psychiatry 2009;80:832–8. [14] Illsinger S, Goken C, Brockmann M, Thiemann I, Bednarczyk J, Schmidt KH, et al. Effect of tacrolimus on energy metabolism in human umbilical endothelial cells. Ann Transplant 2011;16:68–75. [15] Han SY, Chang EJ, Choi HJ, Kwak CS, Suh SI, Bae JH, et al. Effect of tacrolimus on the production of oxygen free radicals in hepatic mitochondria. Transplant Proc 2006; 38:2242–3. [16] Varadhachary AS, Weihl CC, Pestronk A. Mitochondrial pathology in immune and inflammatory myopathies. Curr Opin Rheumatol 2010;22:651–7. [17] Sigal NH, Dumont FJ, Cyclosporin A. FK-506, and rapamycin: pharmacologic probes of lymphocyte signal transduction. Annu Rev Immunol 1992;10:519–60. [18] Hodge S, Hodge G, Ahern J, Liew CL, Hopkins P, Chambers DC, et al. Increased levels of T cell granzyme b in bronchiolitis obliterans syndrome are not suppressed adequately by current immunosuppressive regimens. Clin Exp Immunol 2009;158: 230–6.
Contents lists available at ScienceDirect
Journal of the Neurological Sciences journal homepage: www.elsevier.com/locate/jns
Short communication
Polymyositis in solid organ transplant recipients receiving tacrolimus Gaetano Vattemi a, Matteo Marini a, Marzia Di Chio b, Maria Colpani c, Valeria Guglielmi a, Giuliano Tomelleri a,⁎ a b c
Department of Neurological and Movement Sciences, Section of Clinical Neurology, University of Verona, Italy Department of Medicine and Public Health, Section of Pharmacology, University of Verona, Verona, Italy Department of Gastroenterology, Liver Transplantation Unit, “Ospedali Riuniti”, Bergamo, Italy
a r t i c l e
i n f o
Article history: Received 4 January 2014 Received in revised form 17 June 2014 Accepted 15 July 2014 Available online 6 August 2014 Keywords: Tacrolimus Solid organ transplantation Muscle toxicity Polymyositis Calcineurin inhibitor T cells
a b s t r a c t Tacrolimus, also known as FK506, is an immunosuppressive agent widely used for the prevention of acute allograft rejection in organ transplantation and for the treatment of immunological diseases. This study reports two male patients who underwent solid organ transplantation (liver and kidney). After transplant, the patients received continuous immunosuppressive therapy with oral tacrolimus and later presented clinical manifestations and laboratory signs of myopathy. Muscle biopsies of both patients clearly documented an inflammatory myopathy with the histological features of polymyositis including CD8+ T cells which invaded healthy muscle fibers and expressed granzyme B and perforin, many CD68+ macrophages and MHC class I antigen upregulation on the surface of most fibers. Because of the temporal association while receiving tacrolimus and since other possible causes for myopathy were excluded, the most likely cause of polymyositis in our patients was tacrolimus toxicity. We suggest that patients on tacrolimus should be carefully monitored for serum CK levels and clinical signs of muscle disease. © 2014 Elsevier B.V. All rights reserved.
1. Introduction
2. Case reports
Tacrolimus (FK506) is a calcineurin inhibitor widely used as potent immunosuppressant agent for the prevention of rejection in solid organ transplantation [1,2]. The drug was also introduced in the therapy of several autoimmune diseases including myasthenia gravis and difficult-to-treat cases of polymyositis [3]. In addition, calcineurin inhibitors have been proposed among second-line agents for treatment of myositis and polymyositis, the only “distinctive” neurological manifestations in chronic graft-versus-host diseases (GVHD), a syndrome characterized by immune-mediated multisystemic inflammation occurring after allogeneic hematopoietic stem cell transplantation (HSCT) [4,5]. One study reported a renal transplant patient who had, 10 months after he was started on tacrolimus, an inflammatory myopathy which resolved by its withdrawal [6]. We describe a patient who developed myositis after immunosuppressive treatment with FK506, and performed a detailed analysis of the pathological features of muscle biopsy from this patient and the previous one.
2.1. Patient 1
⁎ Corresponding author at: Department of Neurological and Movement Sciences, Section of Clinical Neurology, University of Verona, P.le L.A. Scuro 10, 37134 Verona, Italy. Tel.: +39 045 8074285; fax: +39.045.8027492. E-mail address: [email protected] (G. Tomelleri).
http://dx.doi.org/10.1016/j.jns.2014.07.036 0022-510X/© 2014 Elsevier B.V. All rights reserved.
This 62-year-old male patient underwent living-donor liver transplantation for alcohol-related cirrhosis two years before. After liver transplant, the patient received continuous immunosuppressive therapy with 1 mg of oral tacrolimus per day. His past medical history disclosed hypertension and diabetes and therapy included low dose of insulin, furosemide, allopurinol and ursodeoxycholic acid. Twenty-one months after the transplantation, the patient developed bile duct stenosis and investigation showed increased levels of serum creatine kinase (CK) (1880 U/L; normal b 200), lactate dehydrogenase (852 U/L; normal b 450), aspartate aminotransferase (160 U/L; normal b 50) and alanine aminotransferase (171 U/L; normal b 50). CK levels were persistently elevated (1600 U/L); electroneurography and needle electromyography documented an axonal peripheral neuropathy. At the time of our observation, three months later, on clinical examination the patient squatted and rose from the floor with difficulty and weakness of the thigh flexor (MRC 4) and quadriceps (MRC 4) muscles was present. Deep tendon reflexes were absent in the arms and legs and vibration sense was lost in the legs. At that time, an open muscle biopsy was obtained from the vastus lateralis. Two months later, the patient underwent surgery for bile duct stenosis but then he developed invasive aspergillosis and two weeks later he died.
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G. Vattemi et al. / Journal of the Neurological Sciences 345 (2014) 239–243
2.2. Patient 2
3.2. Histology and histochemistry
Clinical data have been previously reported [6]. Briefly a 53-year-old man with end-stage renal disease caused by chronic glomerulonephritis underwent a renal transplantation at age 40 and received a second renal graft six years later. A combination treatment of cyclosporine A and azathioprine was started after the second transplant. At age 52 the patient's immunosuppression was switched to tacrolimus, initially 5 mg per day, and within a month 4 mg daily because of high drug serum level. Ten months later the patient complained of muscle pain and laboratory investigations documented a mild increase of CK value, about two–three fold normal value. Because of persistent CK elevations over seven months, an open biopsy of the vastus lateralis muscle was performed. Tacrolimus was then withdrawn followed by resolution of the patient's symptoms and normalization of CK level. The patient was hepatitis C virus (HCV) positive with a viral load of 12.79 MEq/ml two months before muscle biopsy and he was treated with ribavirin which was stopped one month before the elevation of CK.
Serial 8-μm-thick cryosections were stained with standard histological and histochemical methods including hematoxylin and eosin, modified Gomori trichrome, ATPase (pH 4.3, 4.6 and 10.4), succinate dehydrogenase (SDH), cytochrome c oxidase (COX), nicotinamide adenine dinucleotide-tetrazolium reductase (NADH-TR), periodic acid Schiff, Sudan black stains and acid phosphatase.
3. Materials and methods 3.1. Muscle biopsies Muscle biopsies were obtained for diagnostic purposes with written informed consent and the study was approved by the local ethical board.
3.3. Immunohistochemistry and confocal immunofluorescence microscopy Immunohistochemical studies were performed on serial 6.5-μm-thick transverse muscle sections using the following wellcharacterized anti-human antibodies: anti-CD3, anti-CD8, anti-CD20, anti-CD25, anti-CD30, anti-CD57, anti-CD68, anti-MHC (major histocompatibility complex) class I, anti-Granzyme B and anti-Perforin. The reactions were revealed by peroxidase or immunofluorescence methods. Double immunofluorescence was performed using antibodies to CD3 and CD68 in combination with an antibody to CD8. Confocal images were acquired with Zeiss LSM 510 confocal microscope. To control staining specificity the primary antibody was omitted or replaced with nonimmune sera at the same concentration.
Fig. 1. Light microscopy of the muscle biopsies. Histological findings of patient 1 (A–D) and of patient 2 (E and F). A–C (hematoxylin and eosin): Two small endomysial inflammatory infiltrates surrounding and invading healthy muscle fibres (A and B); two necrotic muscle fibers, one of which is entirely invaded by inflammatory cells (B and C); a small basophilic regenerating fiber in C. D: Cytochrome c oxidase shows several COX negative muscle fibres. E, F (hematoxylin and eosin): Three endomysial inflammatory infiltrates surrounding non-necrotic muscle fibers.
G. Vattemi et al. / Journal of the Neurological Sciences 345 (2014) 239–243
4. Results Muscle biopsy from patient 1 documented scattered necrotic fibers invaded by mononuclear cells, basophilic regenerating fibers and small endomysial inflammatory infiltrates invading healthy muscle fibers (Fig. 1 A–C). ATPase reactions showed normal differentiation and distribution of muscle fibers; in addition, sparse lobulated fibers and several COX negative fibers were present by oxidative reactions (Fig. 1D). Inflammatory cells were predominantly composed of CD3 + CD8 + T cells (82% of CD3 + T cells were CD8 +) and CD68 + macrophages (Fig. 2, panels A and B); only single CD20+ B cells were present and CD57+ NK cells were absent. In addition, a few CD25+ T cells and rare CD30+ cells were detected. The cytotoxic CD8+ T cells invading nonnecrotic muscle fibers expressed granzyme B and perforin (Fig. 3A). Muscle fibers invaded by CD8+ T cells showed a sarcolemmal expression of MHC class I antigen, which could be found on several more muscle fibers (Fig. 3B).
241
On muscle biopsy of patient 2 small mononuclear inflammatory cells at endomysial sites were observed; these endomysial inflammatory cells focally surrounded and occasionally invaded non-necrotic muscle fibers (Fig. 1 E,F). The inflammatory infiltrates mainly consisted of CD3 + CD8 + T cells (90% of CD3 + T cells were CD8 +) and CD68 + macrophages (Fig. 2 panel C); neither CD20 + B cells nor CD57 + NK cells were identified. Single CD25 + T cells and CD30 + cells were found. Furthermore, MHC class I antigen was upregulated on the sarcolemma of the majority of the muscle fibers, and granzyme B and perforin were expressed (Fig. 3 C,D). 5. Discussion Tacrolimus is a commonly used T cell-targeted immunosuppressant [1,2]. Adverse effects associated with tacrolimus are mainly dosedependent and include nephrotoxicity, neurotoxicity, hypertension, rhabdomyolysis, altered glucose metabolism, gastrointestinal
Fig. 2. Confocal fluorescence microscopical images. Panel A (patient 1): CD3+ T cells (green) surrounding two non-necrotic muscle fibers are mainly CD8+ T cells (red). Panel B (patient 1): a necrotic muscle fiber invaded by CD68+ macrophages (green) and a healthy fiber surrounded by CD8+ T cells (red). Panel C (patient 2): CD3+ (green) and CD8+ (red) T cells surrounding a non-necrotic muscle fiber.
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G. Vattemi et al. / Journal of the Neurological Sciences 345 (2014) 239–243
Fig. 3. Immunohistochemistry of the muscle biopsies. A, C: Granzyme B staining in muscle biopsy of both patients (A, patient 1 and C, patient 2). B, D: MHC class I antigen is expressed on the sarcolemma of several muscle fibers in both patients (B, patient 1 and D, patient 2).
disturbances, increased susceptibility to infection and malignancy [7]. A recent paper raised the possibility that the drug could induce muscle toxicity [6]. We have report herein two transplant patients who complained of myalgia or muscle weakness, previously described among the side effects of tacrolimus, and had a histologically proven inflammatory myopathy while on immunosuppressive therapy with tacrolimus. The inflammatory myopathy has the features similar to those seen in polymyositis, including CD8 + T cells which invade non-necrotic muscle fibers that express MHC class I antigen [8]. The autoinvasive endomysial CD8+ T cells contain granzyme B and perforin, and MHC class I antigen is ubiquitously upregulated on the surface of most fibers. In view of the temporal association while receiving tacrolimus and since no other cause for myopathy could be determined, we believe that polymyositis in our patients was most likely due to toxicity from tacrolimus. Patient 1 also had clinical and laboratory evidence suggestive of an axonal neuropathy. In our patient diabetes mellitus and chronic alcohol intake were recognized predisposing factors for peripheral neuropathy. In addition, FK506 has been reported to be associated with neuropathy [9]. Although tacrolimus-related neuropathy mainly resembles chronic inflammatory demyelinating polyradiculoneuropathy, patients who developed axonal neuropathy have also been reported [10]. Patient 2 also had HCV infection that in the literature has been reported to be associated with myositis [11]. However, it is unlikely that in our patient the muscle disease was virus-related since the withdrawal of tacrolimus led to the serum CK normalization and the disappearance of muscle symptoms. Skeletal muscle is highly exposed to circulating drugs due to its large global mass and its high metabolic rate and blood flow [12]. Therefore several drugs have been identified as causing toxicity to the muscle tissue [12,13]. Drug-induced inflammatory myopathy has only rarely been
described and offending agents include statins, fibrate, hydroxyurea, penicillamine, omeprazole, niflumic acid, phenytoin, tegafur, interferon alfa, silicon gel, bacilli Calmet-Guerin (BCG) tuberculosis vaccine, alfusozin, the antifungal agent terbinafine [12,13]. At the time of onset of inflammatory myopathy, our patients were not receiving any of all the above drugs. Patient 1 also had histological signs of mitochondrial myopathy suggested by several muscle fibers with COX deficiency. The patient was not treated with any drug inducing mitochondrial toxicity including nucleoside analog reverse transcriptase inhibitors, statins and amiodarone [12,13]. Interestingly, data from previous studies support an adverse effect of tacrolimus on mitochondrial respiratory function [14,15]. On the other hand mitochondrial abnormalities can occur in idiopathic inflammatory myopathies including polymyositis [16]. From the above consideration we reasoned that tacrolimus significantly contributed to the occurrence of inflammatory myopathy in our patients. Paradoxically, an immunosuppressive agent that inhibits T cell activation and proposed for the treatment of refractory polymyositis seems to be related to, or possibly induce T cell-mediated myositis. The mechanism by which tacrolimus might determine deleterious effect on muscle tissue is unknown, but could be comparable to those suggested for other drugs causing an inflammatory myopathy [13]. In addition, tacrolimus prevents programmed cell death of activated T cells, thereby disturbing T cell subset balance and causing an abnormally high number of autoreactive T cells [17]. Moreover, in an in vitro model of T cell activation tacrolimus had only a moderate effect on granzyme B production by CD8+ T cells [18]. We conclude that patients receiving tacrolimus should be made aware of the early muscular symptoms and carefully monitored for CK levels in order to consider reduced dosages of the drug or alternative immunosuppressive therapy.
G. Vattemi et al. / Journal of the Neurological Sciences 345 (2014) 239–243
Funding and conflict of interest statement The authors declare that they have no conflicts of interest, and no funding was required for this work. References [1] Tocci MJ, Matkovich DA, Collier KA, Kwok P, Dumont F, Lin S, et al. The immunosuppressant FK506 selectively inhibits expression of early T cell activation genes. J Immunol 1989;143:718–26. [2] Jang HJ, Kim SC, Han DJ. Tacrolimus for rescue therapy in refractory renal allograft rejection. Transplant Proc 2000;32:1765–6. [3] Schneider-Gold C, Hartung HP, Gold R. Mycophenolate mofetil and tacrolimus: new therapeutic options in neuroimmunological diseases. Muscle Nerve 2006;34:284–91. [4] Wolff D, Schleuning M, von Harsdorf S, Bacher U, Gerbitz A, Stadler M, et al. Consensus conference on clinical practice in chronic GVHD: second-line treatment of chronic graft-versus-host disease. Biol Blood Marrow Transplant 2011;17:1–17. [5] Filipovich AH, Weisdorf D, Pavletic S, Socie G, Wingard JR, Lee SJ, et al. National Institutes of Health consensus development project on criteria for clinical trials in chronic graft-versus-host disease: I. Diagnosis and staging working group report, Biol Blood Marrow Transplant 2005;11:945–56. [6] Orlandi V, Campieri C, Mosconi G, D'Arcangelo GL, Feliciangeli G, Scolari MP, et al. Tacrolimus-associated myositis: a case report in a renal transplant patient. Transplant Proc 2004;36:708–10. [7] Plosker GL, Foster RH. Tacrolimus: a further update of its pharmacology and therapeutic use in the management of organ transplantation. Drugs 2000;59: 323–89.
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[8] Dalakas MC. Review: an update on inflammatory and autoimmune myopathies. Neuropathol Appl Neurobiol 2011;37:226–42. [9] Umapathi T, Chaudhry V. Toxic neuropathy. Curr Opin Neurol 2005;18:574–80. [10] De Weerdt A, Claeys KG, De Jonghe P, Ysebaert D, Chapelle T, Roeyen G, et al. Tacrolimus-related polyneuropathy: case report and review of the literature. Clin Neurol Neurosurg 2008;110:291–4. [11] Ito H, Nagano M, Nakano S, Shigeyoshi Y, Kusaka H. In situ identification of hepatitis C virus RNA in muscle. Neurology 2005;64:1073–5. [12] Klopstock T. Drug-induced myopathies. Curr Opin Neurol 2008;21:590–5. [13] Dalakas MC. Toxic and drug-induced myopathies. J Neurol Neurosurg Psychiatry 2009;80:832–8. [14] Illsinger S, Goken C, Brockmann M, Thiemann I, Bednarczyk J, Schmidt KH, et al. Effect of tacrolimus on energy metabolism in human umbilical endothelial cells. Ann Transplant 2011;16:68–75. [15] Han SY, Chang EJ, Choi HJ, Kwak CS, Suh SI, Bae JH, et al. Effect of tacrolimus on the production of oxygen free radicals in hepatic mitochondria. Transplant Proc 2006; 38:2242–3. [16] Varadhachary AS, Weihl CC, Pestronk A. Mitochondrial pathology in immune and inflammatory myopathies. Curr Opin Rheumatol 2010;22:651–7. [17] Sigal NH, Dumont FJ, Cyclosporin A. FK-506, and rapamycin: pharmacologic probes of lymphocyte signal transduction. Annu Rev Immunol 1992;10:519–60. [18] Hodge S, Hodge G, Ahern J, Liew CL, Hopkins P, Chambers DC, et al. Increased levels of T cell granzyme b in bronchiolitis obliterans syndrome are not suppressed adequately by current immunosuppressive regimens. Clin Exp Immunol 2009;158: 230–6.
