The resurg ence of tuberculosis and the impact of the
study of pulmonary immunopathog enesis*
JOSÉ ROBERTO LAPA E SILVA*, NEIO BOÉCHAT**
The resurgence of tuberculosis as one of the most important infectious diseases to affect mankind came after the illusion
that the disease was under control and would be eradicated before the end of the 20th Century. Over the last 10 years,
in association with American and European research centers, our group at the Universidade Federal do Rio de Janeiro has been dedicated to investigating the pathogenic mechanisms involved in pulmonary tuberculosis. Due to its frequency and role in transmission, pulmonary tuberculosis is the most serious form of the disease. Our hypothesis is that the establishment of latent infection and its progression to active disease depend on an imbalance between activating and deact ivat in g cyt okin es at t he disease sit e. Despit e t he presen ce of prot ect ive mechan isms su ch as t he macrophage expression of phenotypes (denoting cellular and molecular activation of agents involved in protection, such as nitric
oxide and interferon-γ), tuberculosis progresses. A possible explanation for this is the concomitant presence at the site of
infection of molecules such as interleukin- 10 and TGF-β, which are able to deactivate previously activated macrophages.
Recent data suggest that mycobacteria secrete proteins capable of inducing interleukin- 10, thus contributing to overcoming host protective mechanisms. Susceptible individuals would be more able to produce larger amounts of these molecules due to genetic polymorphisms that facilitate interleukin- 10 production at infection onset. The understanding of these mechanisms could advance the prevention and discovery of new therapeutic targets for the control of tuberculosis.
Key words: Tuberculosis/ethiology. Tuberculosis pulmonary/pathology.
St u d y ca rried o u t a t t h e Un iversid a d e Fed era l d o Rio d e J a n eiro (Rio d e J a n eiro Fed era l Un iversit y) a n d Weill Co rn ell Med ica l Co lleg e, Un iversit é d e Pa ris VI (VI Pa ris Un iversit y)**.
Ad d ress fo r co rresp o n d en ce: Un id a d e d e Pesq u isa em Tu b ercu lo se. Av. Brig a d eiro Tro m p o vski, s/ n . Préd io d o HUCFF/ UFRJ , sa la 01 D5 6 , 1 º a n d a r. Ilh a d o Fu n d ã o , Rio d e J a n e iro , RJ CEP 2 1 9 41 - 5 9 0 . Te l: 5 5 2 1 2 5 6 2 2 8 8 7 . E- m a il: jrla p a .n t g @ t e rra .co m .b r
Fin a n cia l su p p o rt : Re d e Bra sile ira d e Pe sq u isa e m TB (Re d e - TB, Bra z ilia n Tu b e rcu lo sis Re se a rch Ne t w o rk)/ Gra n t n o . 6 2 .0 0 5 5 / 01 - 4 - P ACDT-Mile n io .
Abbre viatio ns use d in this pape r:
AIDS – Acq u ire d im m u n o d e ficie n cy syn d ro m e HIV – Hu m a n im m u n o d e ficie n cy viru s HLA- DR – Hu m a n leu ko cyt e a n t ig en - D reg io n M t b – Myco b a ct e riu m t u b e rcu lo sis NOS2 – In d u cib le n it ric o xid e syn t h a se TB – Tu b ercu lo sis
T GF – Tu m o r g ro wt h fa ct o r
INTRODUCTION
Th e p h ra se “t h e resu rg en ce o f…” u sed in o u r t it le is q u it e sig n ifica n t if we co n sid er it a s a m a rket in g p loy. Th o se wh o a re fa m ilia r wit h t h e hist ory of t u bercu losis (TB) in Brazil will recogn ize it a s a n u n t ru t h . In a ct u a lit y, TB h a s n ever cea sed t o b e a p ro b lem in o u r co u n t ry, o r in t h ird - wo rld co u n t ries in g en era l. Wh a t d id ch a n g e wa s t h e p reva len ce o f TB in d evelo p ed co u n t ries. It wa s b elieved t h a t TB h a d b een virt u a lly era d ica t ed in t h e so - ca lled first - wo rld co u n t ries. Pro m in en t a u t h o rs st a t e d t h a t TB wa s b e in g e ra d ica t e d wo rld wid e, just a s sm a llp ox h a d b een . Ho wever, t h ey were a ct u a lly referrin g t o t h e era d ica t io n o f TB in first - wo rld co u n t ries. In t h e 1 9 7 0 s, t h e p reva len ce ra t es o f TB in Ho lla n d were ext rem ely lo w a n d were fa llin g even lo wer. Th e sa m e wa s t ru e in t he rest of Eu rope an d in t he Un it ed St at es. In o t h er wo rd s, first - wo rld co u n t ries were act u ally m a kin g p ro g ress in t h e era d ica t io n o f TB. Wh a t h a s ch a n g ed ? Wh a t h a s m a d e TB a d isea se t h a t is n o lo n g er co n sid ered era d ica b le, a t lea st in t h e b eg in n in g o f t h is n ew cen t u ry? Th e yea r 1 9 8 5 rep resen t s t h e m o m en t at wh ich TB in d ices b eg an t o rise on ce again . This is at t ribu t able t o a n u m ber o f fa ct o rs, o n e o f wh ich is in crea sed m ig ra t io n . Reservoirs of TB exist in Africa, Sou t h Am erica an d Cen t ral Am erica. Asia is o n e o f t h e g reat reservo irs o f TB in t h e wo rld , riva led o n ly b y In d ia in t h e st at ist ics. Con siderin g t hat in t ern at ion al m igrat ion co n t in u e s t o in cre a se , e sp e cia lly f ro m t h e se reservoir areas t o developed cou n t ries, it is obviou s t h a t t h ese reservo irs will even t u a lly b e rep lica t ed in f irst - w o rld c o u n t rie s, d e sp it e TB c o n t ro l p ro g ra m s in t h o se d evelo p ed co u n t ries t h a t o p en t h eir d o o rs t o im m ig rat io n .
Nevert h eless, t h ere is o n e h ist o rical fact o r t h at ca n b e h eld resp o n sib le fo r t h e ch a n g es in t h e TB p reva len ce cu rve: t h e a cq u ired im m u n o d eficien cy syn d ro m e (AIDS) p a n d em ic, wh ich in t ro d u ced a n ew realit y, sin ce in dividu als in fect ed wit h TB who a lso b eco m e in fect ed wit h t h e AIDS viru s ra p id ly co n vert t o a ct ive TB. Ad d it io n a lly, TB d o es n o t re q u ire a ve c t o r f o r it s t ra n sm issio n , b e in g t ra n sm it t e d fro m p e rso n - t o - p e rso n . A h u m a n im m u n odeficien cy viru s- posit ive (HIV+) in dividu al is at great er risk for developin g TB. When t he act ive fo rm o f t h e d isea se is a t t a in ed , t h e b a cillu s m a y b e t ra n sm it t ed t o in d ivid u a ls wh o a re n o t HIV+ , su ch a s h ea lt h p ro fessio n a ls, p riso n m a t es a n d
sh elt er m a t es. Th erefo re, t h e p reva len ce o f TB in creases exp o n en t ially, resu lt in g fro m a sit u at io n t h a t is n o t d irect ly o r n ecessa rily rela t ed t o t h e AIDS viru s. Th is is wh y TB h a s a g a in b een in t h e headlin es an d is con sidered t o be on t he rise, which led t h e Wo rld Healt h Org an izat io n t o d eclare TB a g lo b a l h ea lt h em erg en cy in 1 9 9 5 .
We m u st b ea r in m in d t h a t we a re referrin g t o a d isea se t h a t h a s a fflict ed h u m a n kin d fo r a g es. We kn ow t hat TB has been fou n d in m at erial t aken fro m 4 0 0 0 - yea r- o ld Eg yp t ia n m u m m ies, in wh ich it w a s id e n t if ie d u s in g m o le c u la r b io lo g y t ech n iq u es t h a t em p lo y kn o wn seq u en ces o f t h e gen om e of t hese m ycobact eria. Eviden ce of TB has also been fou n d in t he bon es of Neolit hic hu m an s. In t he Middle Ages, it was called t he “whit e plagu e”. Th ere is a ren ewed in t erest in TB exh ib it ed b y t h e m ed ia , wh ich co u ld resu lt in a d d it io n a l fu n d in g fo r resea rch . In fa ct , la ck o f fin a n cia l su p p o rt fo r r e s e a r c h i s t h e p r i m a r y r e a s o n t h a t o u r u n d erst a n d in g o f TB h a s n o t kep t p a ce wit h t h e a d va n ce o f t h e d isea se. Th erefo re, we view TB a s it wa s in t h e la st cen t u ry. In clin ica l p ra ct ice, we st ill u se t ech n iq u es t h a t h a ve b een u sed fo r u p t o 1 0 0 yea rs. Ma n y o f o u r d ia g n o st ic m et h o d s, su ch a s p u r if ie d p r o t e in d e r iva t ive t e s t in g , w e r e developed 100 years ago. Ou r t herapeu t ic regimen s, in clu d in g a n t i- TB d ru g s su ch a s iso n ia zid a n d st rep t o m ycin , were d evelo p ed 5 0 yea rs a g o . In a d d it io n , t h e va ccin es cu rren t ly in u se, su ch a s t h e b a cillu s Ca lm et t e- Gu érin va ccin e (wh ich wa s d evelo p ed in 1 9 0 9 ) are q u it e d at ed . In vest m en t in TB re se a rch p a le s in co m p a riso n t o t h e so lid fin a n cia l su p p o rt p ro vid ed fo r resea rch in t o o t h er d isea ses su ch a s AIDS.
Bio lo g y a n d Bio m ed icin e, TB h a s effect ively b een left behin d. This is a disease t hat will kill 30 million p eo p le wo rld wid e o ver t h e n ext t en yea rs. Th ere a re few co m p a ra b le sit u a t io n s. Wh en we co n sid er t h e level o f fin a n cia l su p p o rt fo r TB resea rch a n d TB- c o n t ro l p ro g ra m s w o rld w id e , it b e c o m e s obviou s t hat t he su m in vest ed is t ot ally in adequ at e a n d d isp ro p o rt io n a t e. Prio rit y sh o u ld b e g iven t o TB, alt hou gh, u n fort u n at ely, t here are precious few wh o t h in k so .
Even prior t o t he in t rodu ct ion of chemot herapy, e p id e m io lo g ic a l in d ic e s o f TB w e re f a llin g , especially in first - world cou n t ries. At t he begin n in g o f t h e 2 0 t h cen t u ry, t h ere wa s a h ig h p reva len ce o f TB in t h e Un it ed St a t es, wh ere p ro g ressive im p ro vem en t in q u a lit y o f life even t u a lly b ro u g h t abou t a sign ifican t redu ct ion in all epidemiological in dicat ors, even prior t o t he in t rodu ct ion of specific ch em o t h era py. Th e in t ro d u ct io n o f st rep t o m ycin , t he first effect ive an t i- TB dru g, resu lt ed in a secon d co n sid era b le red u ct io n in TB in d ices, wh ich wa s repeat ed years lat er, when ison iazid was in t rodu ced. Th e u n d e r s t a n d in g o f t h e p o s t u la t io n s o f c h e m o t h e r a p y d a t e s f r o m t h e s a m e p e r io d , in d ica t in g t h a t t h e u se o f t h ese d ru g s sh o u ld b e u s e d in c o m b in a t io n in o r d e r t o a vo id t h e appearan ce of resist an t st rain s. From t he begin n in g o f t h e 1 9 5 0 s t o t h e b eg in n in g o f t h e 1 9 8 0 s, t h ere was an en o rm o u s red u ct io n in t h e in cid en ce o f TB in t h e USA, in Eu ro p e a n d in t h e rest o f t h e d evelo p ed wo rld . Ho wever, TB in cid en ce wa s a lso red u ced sig n ifica n t ly in p o o r co u n t ries su ch a s Brazil an d In d ia. Th is was d u e t o an o verall g lo b al im p r o ve m e n t in s a n it a t io n , e ve n in p o o r e r co u n t ries. Gen era l d evelo p m en t , esp ecia lly in t h e a rea s o f t ech n o lo g y, scien ce a n d sa n it a t io n over t h e co u rse o f t h e cen t u ry facilit at ed t h e co n t ro l o f a ll d isea ses. Th is p h en o m en o n is n o t rest rict ed t o TB alo n e, alt h o u g h it s co n t ro l also b en efit ed fro m t he gen eral improvemen t in qu alit y of life. However, midway t hrou gh t he 1980s, t here was an in flect ion in t h e TB p re va le n c e c u rve , re su lt in g in a n a scen d a n t t en d en cy, wh ich st ill p reva ils.
The pathog eny of tuberculosis
The immu n opat hogen y of TB, or rat her t he way t h e h o st im m u n e syst e m a ct s t o im p e d e t h e p ro g ress o f t h e d isea se, is a su b ject t h a t h a s su ffered sig n ifica n t ly fro m t h e la ck o f in vest m en t in research. Amon g t he TB- relat ed fact s men t ion ed
above, on e in part icu lar calls at t en t ion t o t he n eed for research in t his area: TB has in fect ed on e- t hird o f t h e g lo b a l p o p u la t io n , in wh ich t h e re a re a p p ro xim a t e ly 1 0 m illio n ca se s o f a ct ive - TB. Beca u se t h ey p o ssess d efen se m ech a n ism s t h a t keep t h e m yco b act eria in ch eck, t h ereb y im p ed in g it s a ct ivit y, t h e rem a in in g in fect ed in d ivid u a ls d o n o t d evelo p t h e d isea se. Ob vio usly, if we ca n g a in a re a so n a b le u n d e rst a n d in g o f t h is d e f e n se m ech an ism , we will ab le t o co m p reh en d t h e o t h er sid e o f t h e e q u a t io n : wh a t ca u se s 5 % o f a ll co n t a m in a t ed in d ivid u a ls t o d evelo p t h e d isea se.
In fect io n wit h Myco b a ct eriu m t u b ercu lo sis (Mt b ) h a s t h ree p o ssib le o u t co m es: reso lu t io n a t t he poin t of en t ry (du e t o in n at e im m u n it y), act ive d isease o r lat en t TB. In t h e last case, t h e o rg an ism con t rols, bu t does n ot elimin at e, t he in fect ion . The Mt b rem a in s d o rm a n t , rep lica t in g in t erm it t en t ly an d presen t in g an alt ered m et abolism . This creat es a m a ssive reservo ir o f TB. Severa l q u est io n s a re releva n t t o t h is p ro b lem . Wh ich in it ia l even t s lea d t o im m ed ia t e co n t ro l o f t h e in fect io n ? Wh ich fact ors con t ribu t e t o t he on set of lat en t in fect ion ? By id en t ifyin g t h e m ech a n ism s in vo lved , we ca n p r o p o s e va c c in a t io n s , c h e m o t h e r a p ie s a n d a d ju va n t im m u n o t h e ra p ie s t h a t m a y p re ve n t p a t ien t s fro m d evelo p in g t h e a ct ive fo rm o f TB.
Im m u n o lo g y e f f e c t ive ly b e g a n w it h t h e d isco veries m a d e b y Ro b ert Ko ch in t h e 1 8 9 0 s. However, it may be considered a very recen t scien ce, sin ce t he T lym phocyt e an d t he B lym phocyt e were n o t id en t ified u n t il t h e b eg in n in g o f t h e 1 9 7 0 s. Even t h en , we kn ew n o t h in g a b o u t cyt o kin es, t h e u n d e rst a n d in g o f wh ich h a s b ro u g h t a b o u t a revo lu t io n in t h e field o f b io lo g y.
mycobact eria wit hin it s own cyt oplasm. In addit ion t o p ro d u cin g im p o rt an t cyt o kin es, T lym p h o cyt es, in t heir effect or role, represen t a powerfu l weapon , sin ce t h ey a re a b le t o d est ro y m a cro p h a g es t h a t h a ve su ccu m b ed t o t h e m yco b a ct eria . On ce t h e T lym p h o cyt e reco g n izes t h a t t h e m a cro p h a g e h a s lo st t h is b a t t le, it kills t h e m a cro p h a g e, relea sin g t h e b a cillu s in t o t h e en viro n m en t , wh ere m o re e f f e ct ive m a cro p h a g e s ca n p h a g o cyt o se a n d co n t ro l it . Th is rep resen t s a n effect o r a ct io n o f t he T lymphocyt e it self again st mycobact eria, albeit in a n in d irect wa y.
We h a ve b e g u n t o re c o g n iz e c e rt a in c e ll su b p o p u la t io n s, su ch a s t h e T
γ δ
lym p h o cyt es, wh ich p la y a very im p o rt a n t ro le in co m b a t in g TB sin ce t hey are highly t oxic t o in fect ed macrophages an d produ ce cyt okin es t hat st imu lat e t he remain in g m a cro p h a g es t o kill t h e m yco b a ct eria . Even t h e m a cro p h a g e is n o lo n g er sim p ly co n sid ered a n effect o r cell, b u t a lso o n e o f t h e m o st im p o rt a n t in d u cers o f cyt o kin e p ro d u ct io n . In lig h t o f t h is n ew kn owledge, we m oved away from t he con cept t h a t in d u ct io n o f t h e im m u n e resp o n se wo u ld a lwa ys b e ca rried o u t b y T lym p h o cyt es a n d t h a t t h e effect o r a ct io n wo u ld a lwa ys b e p ro vid ed b y t h e m a c r o p h a g e . We n o w kn o w t h a t t h e T lymphocyt e an d t he macrophage may play t he roles o f b o t h in d u ct o rs a n d effect o rs.An o t h er rea so n fo r reeva lu a t in g o u r p revio u s co n cep t io n o f TB is t h e a d ven t o f HIV. Th e AIDS pan dem ic has im posed on hu m an kin d a revolu t ion in t he field on im m u n ology. Never before has t here b een p ro g ress su ch a s t h a t seen in t h e la st 2 0 yea rs. An HIV+ p a t ien t p resen t s a fo rm o f TB t h a t i s v e r y d i f f e r e n t f r o m t h a t s e e n i n im m u n o co m p et en t in d ivid u a ls: it is m u ch m o re severe, m o re o ft en fa t a l a n d h a s a m u ch wo rse effect o n t h e su rviva l o f t h e p a t ien t . In d ivid u a ls d ie m o re ra p id ly wh en su fferin g fro m TB a n d HIV co n co m it a n t ly. Eve n if t h e TB is cu re d , AIDS su rviva l t im e is red u ced(1 ).
The role of cytokines in the development
of tuberculosis
It is essential to understand the mycobactericidal m echan ism s of m acrophages an d how t he im m u n e syst em , especially t he pu lm on ary im m u n e syst em , part icipat es in t hese m echan ism s. In virt u ally all cases, TB t ran sm ission occu rs via t he airways, an d 80% presen t as t he pu lm on ary form of t he disease.
In addit ion , t he pu lm on ary form is t he on ly form t hat is epidem iologically sign ifican t , sin ce it allows person- to- person transmission, thereby maintaining t h e d ise a se t ra n sm issio n cycle . Cyt o kin e s a re horm on es t hat are t he produ ct s of several t ypes of cells wit hin t he organ ism an d have a variet y of fu n ct ions. In t he case of TB, specifically, t hey play very im port an t roles, which we are on ly begin n in g t o u n derst an d.
We u n d erst a n d t h a t , t o b e p ro t ect ed a g a in st TB, an in dividu al n eeds t o possess a part icu lar t ype o f cyt o kin e t h a t en a b les m a cro p h a g es t o kill t h e m yco b a ct eria . Ho wever, we a re a lso b eg in n in g t o u n derst an d t hat t here are cert ain t ypes of cyt okin es t h a t , in versely, d ea ct iva t e m a cro p h a g es, t h ereby p reven t in g t h e d est ru ct io n o f t h e m yco b a ct eria . Th ere a re cyt o kin es o f b o t h Th 1 a n d Th 2 p ro files. In sh o rt , t h e Th 1 cyt o kin es are t h o se, su ch as
IFN-γ
a n d IL- 2 , w h ich a ct iva t e t h e m a cro p h a g e m icrobicidal m echan ism s, an d Th2 cyt okin es, su ch as IL- 4, IL- 5 an d IL- 10, deact ivat e t he macrophage. Dep en d in g o n t h e p reva len ce o f t h ese t yp es o f cyt o kin es, TB will p ro g ress o r will b e p reven t ed . Ou r h yp o t h esis is t h a t t h ere is a p reva len ce o f d eact ivat in g cyt o kin es wh en t h e d isease d evelo p s, even wh en a ct iva t in g cyt o kin es a re p resen t . If t h e in fect ed in d ivid u a l h a s a g en et ic t en d en cy t o p ro d u ce a g rea t er a m o u n t o f IL- 1 0 , fo r exa m p le, h is ch a n ce s o f d e ve lo p in g TB will b e h ig h e r. Therefore, gen et ic polym orphism s, which facilit at e a g rea t er p ro d u ct io n o f IL- 1 0 wh en fa cin g a n in fect io u s d isease, wit h in cert ain p o p u lat io n s m ay b e u sed in t h e fu t u re a s g en et ic m a rkers fo r t h e st u d y o f g ro u p s wit h h ig h e r o r lo we r risk o f d evelo p in g a ct ive TB. Th ere a re wa ys t o id en t ify popu lat ion grou ps which are high or low produ cers o f IL- 1 0 . If a n in d ivid u a l c o n t a m in a t e d b y m yco b act eria is a m ajo r p ro d u cer o f IL- 1 0 , h e will b e t rea t ed wit h g rea t er ca re, a n d will p ro b a b ly b e req u ired t o t a ke a n IL- 10 in h ib it o r.dem on st rat ed t hat wild- t ype m ice, t hat produ ce a lower amou n t of IL- 10, su rvive the same dose levels, sh o win g t h a t exa g g era t ed p ro d u ct io n o f IL- 1 0 redu ces t he chan ce of su rvivin g a m ycobact erial in fect ion .
Ou r g ro u p a t t h e Un iversid ad e Fed eral d o Rio d e J an eiro (Rio d e J an eiro Fed eral Un iversit y) h as wo rked in co n ju n ct io n wit h Co rn ell Un iversit y in New Yo rk fo r t h e la st t en yea rs, d evelo p in g a n d pu blishin g variou s st u dies t hat focu s on evalu at in g cells collect ed from TB- affect ed areas of t he lu n gs. Ou r a im h a s b een t o d et erm in e wh et h er t h ere is en h a n ced o r red u ced exp ressio n o f IL- 1 0 o r o t h er cyt o kin es wit h sim ilar act io n s t h at co u ld g en erat e t h is im m u n o lo g ica l d ysf u n ct io n . Ou r st u d ie s in clu ded pat ien t s wit h pu lm on ary TB su bm it t ed t o d ia g n o st ic b ro n ch o a lveo la r la va g e b eca u se t h ey were n o t p ro d u cin g sp u t u m o r p ro d u ced sp u t u m t h a t t e s t e d n e g a t ive f o r m yc o b a c t e r ia . Th e t ech n iq u e in vo lves d ra win g a n d cen t rifu g in g t h e flu id , a ft er wh ich t h e su p ern a t a n t is st o red a n d t he cells are st u died. In order t o det ermin e whet her t h e r e is e x a g g e r a t e d e x p r e s s io n o f c e r t a in cyt o kin es, t h e wh o le cell m a y b e st u d ied in t h e cyt o ce n t rif u g e d m a t e ria l, o r t h e ce ll ca n b e flat t en ed an d t he gen et ic mat erial ext ract ed. In on e o f t h e e a r ly s t u d ie s(3 ), w e d e t e r m in e d t h e
phen ot ype of t he T lymphocyt es an d macrophages, a n d we fo u n d t h a t TB p a t ien t s wit h co n co m it a n t HIV a ls o p r e s e n t e d d ys f u n c t io n w it h in t h e m a cro p h a g e p o p u la t io n . In g en era l, we b elieve t h a t , in HIV in fect io n , t h ere a re b a sic d efect s in t h e T CD4 + lym p h o cyt e, wh ereas, in p at ien t s wit h TB an d HIV, t he macrophage it self becomes u n able t o g en era t e a d efen se a g a in st t h e m yco b a ct eria . Ep it h elio id cells id en t ified b y sp ecific m o n o clo n al a n t ib o d y a re p re se n t in sig n if ica n t ly g re a t e r n u m b ers in t h e b ro n ch o a lveo la r la va g e flu id o f immu n ocompet en t pat ien t s, whereas t heir n u mbers a re red u ced in HIV+ p a t ien t s. We a lso st u d ied t h e m acro p h ag e exp ressio n o f cert ain m o lecu les, su ch a s t h e h u m a n leu ko cyt e a n t ig en - D reg io n (HLA-DR), wh ich is a T lym p h o cyt e a n t ig en - p resen t in g c e ll. Wh e n t h is m o le c u le is f o u n d in g r e a t q u a n t it ie s , it r e s u lt s in e n h a n c e d a n t ig e n -p resen t in g a b ilit y. In TB -p a t ien t s, t h e q u a n t it y o f HLA- DR is m u ch g reat er t h an in co n t ro ls, wh ereas in TB pat ien t s wit h con comit an t HIV it is very mu ch d im in ish ed , d em o n st ra t in g a d efect wit h in t h e m acro p h ag e it self.
We a lso st u d ied n it ric o xid e, wh ich is a n o t h er m o lecu le t h a t is very im p o rt a n t in t h e d efen se again st mycobact eria, an d t he n it ric oxide in du ct or e n z ym e , a lso kn o wn a s in d u cib le n it ric o xid e syn t h a se (NOS2 ). We d e m o n st ra t e d in cre a se d en zym e exp ressio n(4 ) wit h in t h e p o p u la t io n o f
p a t ien t s wit h TB. Th erefo re, a lt h o u g h we h a ve dem on st rat ed t hat t here is an in crease of HLA- DR, a s well a s a n in crea se in t h e NOS2 en zym e (b o t h o f wh ich sh o u ld , t h e o re t ica lly, p ro t e ct t h e se in d ivid u a ls), t h ey co n t in u e t o b e ill. We b elieve t h is is b eca u se, even in t h e p resen ce o f in crea sed exp ressio n o f t h ese d efen se fa ct o rs, so m et h in g is im p ed in g t h e p ro t ect io n . We h yp o t h esize t h at t h e d ea ct iva t in g cyt o kin es a re resp o n sib le fo r t h is h in d ran ce. In a su b seq u en t st u d y, we assessed t h e q u a n t i t y o f c y t o k i n e s p r e s e n t i n t h e b ro n ch o a lveo la r la va g e flu id o f TB p a t ien t s(5 ). We
det ermin ed t hat levels of t he cyt okin e t hat act ivat es macrophage microbicidal fu n ct ion , n amely IFN-
γ ,
a re in crea sed , a s we wa n t t h em t o b e. Ho wever, t h e p a t ien t p resen t s a ct ive TB a n d will d ie if n o t t rea t ed , d em o n st ra t in g t h a t IFN-
γ
a lo n e h a s n o effect . This is probably du e t o t he fact t hat , wit hin t h e sa m e m a t e ria l, t h e re is c o e xp re ssio n o f deact ivat in g cyt okin es, which perform a sign ifican t p h ysio lo g ica l fu n ct io n in co n t a in in g t h e im m u n e rea ct io n s a ft er t h e st im u lu s fo r t h a t rea ct io n h a s b een wit h d ra wn . Th ese a re a n t i- in fla m m a t o ry cyt o kin es, wh ich are im p o rt an t in in t erru p t in g t h e in fla m m a t o ry p ro cess. Th e p ro b lem is t h a t t h eir exp ressio n is a p p a ren t ly in crea sed wh en it sh o u ld n o t b e. Recen t st u d ies co n d u ct ed b y o u r g ro u p h a ve sh o wn t h is co exp ressio n o f a ct iva t in g a n d d e a ct iva t in g cyt o kin e s. Fu rt h e rm o re , a n o t h e r d ea ct iva t in g cyt o kin e kn o wn a s t u m o r g ro wt h fact or-β
(TGF-β )
is also in creased. In pat ien t s wit h TB, we h a ve d em o n st ra t ed t h a t , in a d d it io n t o in crea sed exp ressio n o f t h is cyt o kin e, n u m b ers o f t h e cellu la r recep t o rs req u ired fo r TGF-β
a ct ivit y a re a lso in crea sed(6 ).o f sig n if ica n t ly in t e rf e rin g wit h t h e im m u n e re sp o n se – wo rkin g in it s fa vo r, so t o sp e a k. Recen t ly pu blished st u dies con du ct ed by ou r grou p at t he Un iversit y of Corn ell show t hat on e of t hese p ro t ein s, kn o wn a s CFP3 2 , is exp ressed o n ly b y m em b ers o f t h e Mt b co m p lex an d is n o t exp ressed b y o t h er sp ecies o f m yco b a ct eria . In vit ro an d ex vivo st u d ies h ave sh o wn t h at t h is p ro t ein is ab le t o in du ce con siderable produ ct ion of IL- 10, which, a s p revio u sly m en t io n ed , fa cilit a t es t h e p ro g ress o f t h e in fect io n(7 ). Th e p resen ce o f t h is g en e in
t h e m yco b a ct eria h a s a d va n ced t h e d evelo p m en t o f n ew d ia g n o st ic m et h o d s a s well a s a id in g t h e iden t ificat ion of su bspecies wit hin t he gen u s, wit h im p lica t io n s fo r st u d ies t h a t fo cu s o n m o lecu la r ep id em io lo g y(8 ).
Role of dendritic cells
An ot her cell t ype t hat m erit s fu rt her st u dy, du e t o it s a b ilit y t o in d u ce a p o t en t ia lly lo n g - la st in g prot ect ive im m u n e respon se t o TB, is t he den drit ic ce ll. Su fficie n t n u m b e rs o f t h is t yp e o f ce ll, t oget her wit h ot her t ypes of cells of t he mon ocyt e-m a cro p h a g e lin ea g e, a re essen t ia l fo r g ra n u lo e-m a form at ion , which sim u lt an eou sly prot ect s t he host fro m d issem in a t io n o f t h e in fect io n a n d p ro t ect s the microorgan ism from bein g elimin ated(9). A den se
cellu la r p o p u la t io n a n d t h e p resen ce o f cert a in cyt o kin es, su ch a s t yp e- I in t erfero n , a re essen t ia l fo r g ra n u lo m a fo rm a t io n a n d red u ct io n o f t h e b a ct eria l lo a d(1 0 ). Recen t st u d ies co n d u ct ed b y o u r
g ro u p , in a sso cia t io n wit h resea rch ers fro m t h e In st it u t Past eu r d e Paris (Paris Past eu r In st it u t e) h a ve sh o wn t h e im p o rt a n ce o f t h ese m ech a n ism s in est a b lish in g a p ro t ect ive resp o nse t o TB(11 ,1 2 ).
In co n clu sio n , t h e st u d y o f t h e m ech a n ism s t h ro u g h wh ich t h e h o st d efen d s it self ag ain st Mt b in fect io n , a s well a s o f t h e p o t en t ia l m ech a n ism s b y wh ich t h e m yco b a ct eria m a n ip u la t es t h e h o st im m u n e syst em in it s o wn fa vo r, m a y sh ed n ew lig h t o n t h e p a t h o g en y o f TB. Th e resu lt o f t h is wo u ld b e b et t er va ccin es, n ew t h era p eu t ic t a rg et s a n d m o r e p r e c is e d ia g n o s t ic m e t h o d s f o r co m b at in g t h is t errib le illn ess.
REFERENCES
1 . Krit ski AL, La p a e Silva J R, Co n d e MB. Tu b e rcu lo sis a n d HIV: ren ewed ch a llen g e. Mem . In st . Oswa ld o Cru z 1 9 9 8 ; 9 3 : 41 7 - 2 1 .
2 . Murray PJ, Wang L, Onufryk C, Tepper RI, Young RA. T cell-d e rive cell-d IL- 1 0 a n t a g o n iz e s m a c ro p h a g e f u n c t io n in mycobact erial in fect ion . J . Im m u n ol. 1997;158:315- 21. 3 . La p a e Silva J R, Lin h a res C, Bo ech a t N, Reg o L, Bo n ecin i
Alm e id a MG, Krit ski AL, e t a l. P h e n o t yp e s o f lu n g m o n o n u clea r p h a g o cyt es in t h e b ro n ch o a lveo la r la va g e f l u i d o f H IV s e r o n e g a t i v e t u b e r c u l o s i s p a t i e n t s : evid en ce fo r n ew recru it m en t a n d cell a ct iva t io n . Mem . In st . Oswa ld o Cru z 1 9 9 6 ; 91 :3 8 9 - 9 4 .
4 . Nich o lso n S, Bo n e cin i- Alm e id a MG, La p a e Silva J R, Na t h a n C, Xie QW, Mu m f o rd R, We id n e r J R, e t a l. In d u cib le n it ric o xid e syn t h a se in p u lm o n a ry a lveo la r m a cro p h a g es fro m p a t ien t s wit h t u b ercu lo sis. J . Exp . Me d . 1 9 9 6 ; 1 8 3 : 2 2 9 3 - 3 0 2 .
5 . Bo n e c in i- Alm e id a MG, La p a e Silva J R, Krit ski AL, Mo rg a d o MG, Na t h a n C, Ho J L Im m u n e resp o n se d u rin g HIV a n d t u b ercu lo sis co - in fect io n . Mem . In st . Oswa ld o Cru z 1 9 9 8 ; 9 3 : 3 9 9 - 4 0 2 .
6 . Bon ecin i Almeida MG, HoJL, Boéchat N, Hu ard RC, Chit ale S, Do o H, e t a l.Do wn - Mo d u la t io n o f Lu n g Im m u n e Resp o n ses b y In t erleu kin - 1 0 a n d Tra n sfo rm in g Gro wt h Fact or-β (TGF-β) an d An alysis of TGF-β Recept ors I an d II in Act ive Tu bercu losis. In fect Immu n 2004; 72:2628- 34. 7 . Hu a rd RC, Ch it a le S, Leu n g M, La zza rin i LCO, Zh u H, Sh a sh kim a E, e t a l.Th e Myco b a ct e riu m t u b e rcu lo sis
co m p le x- re st rict e d g e n e cp f3 2 (Rv0 5 7 7 ) e n co d e s a n e x p r e s s e d p r o t e i n t h a t i s d e t e c t a b l e i n h u m a n t u b ercu lo sis p at ien t s. In f. Im m u n . 2 0 0 3 ; 71 :6 8 71 - 6 8 8 3 . 8 . Hu ard RC, Lazzarin i LCO, Bu t ler WR, van Sooligen D, Ho JL. A PCR- base met hod t o differen t iat e t he su bspecies of t he Mycobact eriu m t u bercu losis complex on t he basis of g en o m ic d elet io n . J Clin Microb. 2 0 0 3 ; 41 :1 6 3 7 - 5 0 . 9 . Bo ech at N, Bo u ch o n n et F, Bo n ay M, Gro d et A, Pelicic V,
Gicq u el B, et a l. Cu lt u re a t h ig h d en sit y im p ro ves t h e ab ilit y o f h u m an m acro p h ag es t o co n t ro l m yco b act erial g ro wt h . J . Im m u n o l. 2 0 01 ; 1 6 6 :6 2 0 3 - 11 .
1 0 . Bo u ch o n n et F, Bo ech a t N, Bo n a y M, Ha n ce AJ . Alp h a / b e t a i n t e r f e r o n i m p a i r s t h e a b i l i t y o f h u m a n m a c ro p h a g e s t o c o n t ro l g ro w t h o f Myc o b a c t e riu m b o vis BCG. In fect Im m u n 2 0 0 2 ; 7 0 :3 0 2 0 - 5 .
1 1 . Na h o ri M- A, La g ra n d erie M, Lefo rt J , Th o u ro n F, J o sep h D, Win t e r N, e t a l. Ef f e ct s o f Myco b a ct e riu m b o vis
BCG o n t h e d evelo p m en t o f a llerg ic in fla m m a t io n a n d b ro n ch ia l h yp erresp o n siven ess in h yp er- Ig E BP2 m ice va ccin a t ed a s n ewb o rn s. Va ccin e 2 0 01 ; 1 9 : 1 4 8 4 - 9 5 . 1 2 . La g ra n d erie M, Na h o ri MA, Ba la zu c AM, Kiefer- Bia sizzo
