D E N G U E

MUSCLE BIOPS Y FINDING S I N 1 5 PATIENT S

S.M.F. MALHEIROS* , A.S.B . OLIVEIRA* , B . SCHMIDT 1** J.G . CAMARG O LIMA* , A.A. GABBAI *

SUMMARY — Dengu e i s know n t o produc e a syndrom e involvin g muscles , tendon s an d joints . The hallmar k o f thi s syndrom e i s sever e myalgi a bu t include s fever , cutaneou s rash , an d headache. Th e neuromuscula r aspect s o f thi s infectio n ar e outline d onl y i n isolate d reports , and th e muscl e histopathologica l feature s durin g myalgi a hav e no t bee n described . I n orde r to ascertai n th e actua l neuromuscula r involvemen t in dengu e an d bette r comprehen d th e histological natur e o f myalgia , w e performe d a clinica l an d neurologica l evaluation , a seru m CPK leve l an d a muscl e biops y (wit h histochemistry ) i n 1 5 patient s ( 4 males) , media n ag e 23 year s (rang e 14-47 ) wit h classi c dengu e fever , serologicall y confirmed , durin g th e bra -zilian dengu e epidemic s fro m Septembe r 198 6 t o Marc h 1987 . Al l patient s ha d a histor y o f fever, headach e an d sever e myalgia . Upo n examinatio n 4 ha d a cutaneou s rash , 3 ha d fever , and 3 a smal l hepatomegaly . Th e neurologica l examinatio n wa s unremarkabl e i n al l an d included a manua l muscl e test . CP K wa s mildl y elevate d i n onl y 3 patients i Muscl e biops y revealed a ligh t t o moderat e perivascula r mononuclea r infiltrat e i n 1 2 patient s an d lipi d ac -cumulation i n 11 . Mil d mitochondria l proliferatio n wa s see n i n 3 , fe w centra l nucle i i n 3 , rare foc i o f myonecrosi s i n 3 , an d 2 patient s ha d typ e grouping . Dengu e i n ou r patients , produced myalgi a bu t n o detectabl e muscl e weaknes s o r othe r neuromuscula r involvement . The mai n histopathologica l correlatio n wit h myalgi a seem s t o b e a perivascula r mononuclea r infiltrate an d lipi d accumulation .

KEY WORDS : dengue , muscle , myalgia .

Dengue: achado s d e biópsi a muscula r e m 1 5 paciente s

RESUMO — A síndrom e clínic a produzid a pel o víru s d a dengue , inclu i febre , exantema , cefaléia e especialment e mialgia . Entretanto , a s possívei s alteraçõe s morfológica s d o muscul o es -quelético, eventualment e relacionada s co m a mialgia , aind a nã o havia m sid o estudada s e m sere s humanos co m dengue » Noss o objetiv o fo i estuda r o substrato , anatomopatológic o d a mialgi a nesses pacientes . Fora m avaliado s 1 5 paciente s co m diagnóstic o d e dengue , form a clássica , co m idades variand o d e 1 4 a 4 7 anos (median a d e 2 3 anos) , send o 4 d o sex o masculin o e 1 1 do sex o feminino, atravé s d e exam e clínic o e neurológico , exame s laboratoriai s e biópsi a muscula r co m histoquímica, durant e a epidemi a d e dengu e e m Alagoas , e m 1987 . Todos os paciente s apresenta -vam históri a d e cefaléia , febr e e mialgi a intensa , se m fraquez a muscular . A o exam e clínic o observou-se exantem a e m 4 pacientes , febr e e m 3 e discret a hepatomegali a e m 3 . O exam e neurológico fo i norma l e m todo s e a enzim a C K s é ri ca estav a pouc o elevad a e m 3 pacientes . A biópsi a muscula r revelo u discret o infiltrad o inflamatóri o mononuclea r perivascula r e m 1 2 pacientes, acúmul o lipídic o e m 11 , predominânci a d e fibra s d o tip o I e m 6 , raro s foco s d e necrose e m 3 , proliferaçã o mitocondria l e m 3 , centralizaçã o nuclea r e m 3 e "typ e grouping " em 2 . A s alteraçõe s mai s frequentement e observadas n a biópsi a muscular , infiltrad o inflama -tório perivascula r e acúmul o lipídico , pode m esta r relacionada s co m a mialgia .

PALAVRAS-CHAVE: dengue , músculo , mialgia .

•Department o f Neurolog y an d **Dep»artmen t o f Pathology , Escol a Paulist a d e Medicina , São Paulo . Aceite : 08-novembro-1992 .

Dr. Acar y S.B . Oliveir a — Departmen t o f Neurology , Escol a Paulist a d e Medicin a — Ru a Botucatu 72 0 — 04023-900 São Paul o S P — Brasil.

Dengue, caused by an arthropod-borne RNA virus i s stil l a major public health proble m i n tropica l areas , mainl y i n Southeas t Asia , th e Pacifi c region , and mor e recently , i n th e America s 5,8,22. Myalgi a i s a characteristi c sympto m of dengu e fever , an d i s als o frequen t i n othe r vira l infection s 14,16,1 8 bu t it s pathogenesis stil l remains poorly understood 14,16. Ther e are isolated experimenta l reports abou t th e pathologica l finding s i n skeleta l muscl e i n dengu e L2**, bu t we have been unable to fin d an y pathological stud y of skeleta l muscl e i n humans with myalgi a an d dengu e fever .

This repor t describe s th e skeleta l muscl e histologica l abnormalitie s i n 1 5 patients wit h dengu e feve r an d sever e myalgia , a s a n attemp t t o bette r unders * tand th e histologica l natur e o f tha t symptom .

PATIENTS AN D METHODS

We studie d 1 5 patient s wit h dengu e fever , 4 males , 1 1 females , media n ag e 2 3 year s (range 1 4 t o 4 7 years) , durin g a n epidemi c o f classi c dengu e feve r fro m Septembe r 198 8 t o March 1987 , i n th e cit y o f Maceio , stat e o f Alagoas , Brazil . Th e diagnosi s wa s base d o n epidemiological data , clinica l manifestation s an d confirme d b y hemagglutinatio n inhibitio n method (positiv e in al l 1 5 patients) , Ig M antibod y detectio n b y Ma c EDIS A (positiv e i n al l 8 studied ) an d viru s isolatio n (i n on e patien t studied) .

All patient s ha d a histor y o f fever , headach e an d sever e myalgia . Tw o patient s complain -ed als o o f arthralgia . Al l patient s wer e submitte d t o clinica l an d neurologica l examination s including a manua l muscl e tes t 2. Tabl e 1 summarize s th e age , sex , clinica l signs , neurologica l examination an d duratio n o f symptom s befor e biopsy . Classi c dengu e feve r i s typicall y a n ucute febril e illnes s wit h rash , headache , retro-orbita l pain , arthralgi a an d sever e myalgi a 13,21.

Blood test s tha t include d complet e bloo d count , sodium , potassium , lacti c dehydroge -nase, creatin e kinas e (CK) , pyruvi c transaminas e (SGPT) , an d glutami c transaminas e (SOOT ) were unremarkabl e wit h th e exceptio n o f thre e patient s (patient s 4 , 7 an d 13 ) tha t showe d a fligh t increas e o f seru m C K ( 2 time s normal) . Urinalysi s an d EK G wer e norma l i n al l patients. Electromyograph y wa s no t done .

Muscle biopsie s wer e performe d i n th e lef t superficia l deltoi d muscl e o n al l patients . The biops y techniqu e an d muscl e processin g use d ar e al l wel l establishe d an d hav e bee n described elsewher e 9 . Al l biopsie s wer e staine d wit h H&E , modifie d Gomori , PAS' , Suda n red, ATPase s 9.4 , 4. 3 an d 4.6 , NAD H an d SDH .

RESULTS

All biopsie s presente d norma l stainin g t o PA S an d NADH . Th e histopathologica l findings ar e summarize d i n Tabl e 2 .

A ligh t t o moderat e perivascula r mononuclea r infiltrat e wa s foun d i n 1 2 patient s (H&E ) (Fig. 1) . Th e infiltrat e consiste d o f mononuclea r cell s aroun d smal l vessel s mainl y i n th e perimysium. Th e infiltrat e wa s predominan t i n th e perivascula r spac e an d di d no t invad e miuttle fibers . I n rar e occasion s th e infiltrat e was als o see n i n th e endomysiu m mainl y around smal l vessel s (Fig . 2. ) Eve n whe n moderat e th e infiltrat e di d no t invad e o r disrup t the vesse l walls . Th e infiltrat e wa s foun d i n 1 1 patient s withi n 2 t o 8 day s o f symptoma -tology; onl y on e patien t ha d a 1 4 da y histor y o f dengu e fever .

Eleven biopsie s presente d wit h a mil d lipi d accumulatio n (Suda n red ) diffusel y locate d in a t leas t 50 % o f th e fibers . Nin e biopsie s showe d bot h perivascula r infiltrate s an d lipi d accumulation.

Other finding s include d mitochondria l proliferatio n locate d mainl y i n th e peripher y o f the fiber s i n 3 biopsies , centra l nucle i i n 3 , rar e foc i o f necrosi s i n 3 , an d typ e groupin g in 2 . Thes e latte r finding s wer e alway s associate d wit h eithe r perivascula r infiltrates , lipi d accumulation o r bot h (Tabl e 2).

Except fo r on e patien t (wit h a 3 da y history ) tha t presente d a norma l muscl e biopsy , 11 patient s showe d a t leas t tw o different abnormalitie s i n thei r biopsies .

COMMENTS

To ou r knowledg e thi s i s th e firs t repor t o f th e muscl e histopathologica l features tha t accompan y classi c dengu e fever . I t i s intriguin g t o not e tha t w e have not foun d any exampl e o f tru e myositi s i n spit e o f th e fac t tha t myositis , sometimes associate d wit h perivascula r inflammatory infiltrates, seem s t o b e the basic histologica l abnormalit y found i n skeleta l muscl e o f animal s an d humans infected wit h othe r viruse s lik e influenz a an d coxsacki e 11.12,17.

Agarawal e t al. 1 hav e studied metaboli c change s i n skeleta l muscl e i n mice f̂ter intracerebra l inoculatio n o f Dengu e virus typ e 2 but d o not describ e histo -

logical muscl e abnormalities . Nat h e t al.2 0 hav e reporte d th e ultrastruetura l alterations i n skeleta l muscl e o f dengu e infecte d mic e showin g destructio n o f myofibrils an d sarcoplasmi c reticulum , swellin g o f mitochondria , glycoge n accu -mulation an d th e demonstratio n o f virus-lik e particle s i n th e perinuclea r zon e and cytoplasm . The y als o d o no t describ e an y histologica l ligh t microscop y abnormalities.

Our finding s showe d perivascular mononuclear infiltrates (bu t no myositis) and lipi d accumulatio n (bu t no t glycogen) . Perivascula r infiltrate s wer e des -cribed i n th e CNS7 , sof t tissue 4 an d othe r organs 2! i n pos t morte m studie s of patient s wit h dengu e shoc k syndrom e but no t i n th e ski n i n dengu e hemor-rhagic feve r 24.

The perivascula r infiltrat e foun d i n 1 2 of ou r 1 5 patients ma y be relate d to the pathogenesis o f myalgia by producing ischaemic o r metabolic changes 1.16,23 or eve n b y loca l releas e o f short-actin g chemica l mediator s tha t coul d likel y b e effectors o f pain 16,23. W e found lipid accumulation in 1 1 patients an d 9 of the m also showe d perivascular infiltrates . Th e lipid accumulatio n coul d be th e conse -quence o f a n ischaemi c phenomeno n relate d t o th e perivascula r abnormality .

The mitochondria l proliferatio n foun d i n thre e o f ou r patient s an d th e mitochondrial change s foun d b y Nat h e t al. 2^ in thei r ultrastructura l stud y o f skeletal muscl e i n mic e ma y als o reflec t a n abnormalit y i n th e oxidativ e meta -bolic pathway and might suggest tha t myalgi a is du e to metaboli c change s 1.16,23.

Three of ou r patients wh o presented wit h perivascula r infiltrate s an d lipid accumulation, als o showe d som e foc i o f necrosis . Althoug h these patient s di d not complai n o f a mor e sever e myalgi a tha n th e others , muscl e necrosi s i s known to produce pain 16,23. On e of thes e patient s als o showe d a mildly elevated serum CK . Henc e i t i s possibl e tha t th e othe r tw o patient s wit h elevate d CK and th e other s withou t necrosi s an d C K abnormalities ma y hav e a metaboli c muscle involvemen t sever e enoug h t o produc e pai n bu t no t enoug h t o produc e necrosis o r C K elevation.

As a n indirec t evidenc e o f altere d sarcoplasmi c homeostasis , possibl y als o reflecting th e metabolic abnormalities already cited, three o f ou r patients showed central nucle i 10.

We hav e bee n unabl e t o explai n th e reaso n fo r th e typ e groupin g foun d in 2 of ou r 15 patients. I n the superficia l deltoi d muscl e th e ATPase-checkboard pattern rarel y varie s 345 an d thes e patient s di d no t sho w an y clinica l evidenc e of periphera l nerv e involvement .

New studie s ar e neede d t o analyz e th e vascula r involvemen t i n muscl e in th e genesi s o f myalgia . Immunohistochemica l an d i n sit u hybridizatio n tech-niques wil l hel p defin e th e presenc e o f th e viru s i n cell s aroun d vessel s an d in th e muscl e an d determine th e natur e o f th e mononuclea r cells involved.

Aknowledgements — W e ar e gratefu l t o Dr . Fernand o Lir a Net o fo r referrin g th e patients t o u s an d fo r hi s assistanc e durin g ou r sta y i n Maceió , t o Institut o Adolf o Lut z (São Paulo , SP , Brasil ) fo r performin g th e Ig M antibod y detection s b y Ma c ELIS A an d the virus * isolation , an d t o Institut o Evandr o Chaga s (Belém , PA , Brasil ) fo r performin g the hemagglutinatio n inhibitio n tests .

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