PENGARUH PEMBERIAN L-ARGININ TERHADAP RESPON IMUNITAS SELULER MENCIT BALB/c YANG DIINOKULASI SALMONELLA TYPHIMURIUM

Abstract

Background: Salmonella typhi infection can suppress cellular mediated immunity. Approriate therapeutic strategy is needed to improve this condition, by means of combining antibiotic and immunostimulatory agent. L-Arginine, an amino acid as NO precursor, can act as immunomodulator, through GH and polyamine. Objective: This research was aimed to examine effects of L-Arginine on cellular mediated immunity of BALB/c mice were inoculated with Salmonella typhimurium. Method: An animal experimental study was carried out, using the post test only control group design in BALB/c mice. A total of 102 healthy male mice, 8-10 weeks, 30-40 g then divided randomly, 42 mice divided into 7 group. The first group K(-) was without treatment; second group K(+) was inoculated with Salmonella typhimurium; the third P1 was given L-arginine HC1 1,2% 7 days in drinking water, and without infection; the fourth P2 was given L-arg.inine HC1 1,2% 7 days in drinking water as continuous preatreatment then on day 8 was inoculated with Salmonella typhimurium; the fifth P3 as intermitten type pretreatment was given L-arginine HC1 1,2% 7 days intermitten in drinking water then was inoculated with Salmonella typhimurium; the sixth P4 as pre/posttreatment was given L-arginine HC1 1,2% 7 days before infection and continued until sacrified. Mice were sacrified on day 3rd of infection. The seventh P5 as postreatment was given L-arginine HO 1,2% 7 days after infection, then were sacrified. The dosage of Salmonella typhimurium was 106. After being sacrified, then was examined lymphocyte proliferation response, phagocytosis by using latex beads, NO production in plasma and supernatant of cultured peritoneal macrophage by the Griess method, and ROI production was measured in phorbol myristate acetate-stimulated macrophage (NBT method), and bacterial growth. Survival examination used 60 mice divided 5 group K(+), P2, P3, P4, P5 for 21 days. It was used 107 Salmonella typhimurium. Cellular immunity were analyzed by using ANOVA and Kruskal Wallis test. Survival rate was determined by using Kaplan Meier method and analyzed by using Logrank test Result: The results showed that L-arginine as continuous pretreatment, intermitten type pretreatment dan pre/posttreatment significantly increased the count of lymphocyte (p=0,037; p=0,037; p=0,010). L-arginine as continuous pretreatment, intermitten type pretreatment ,dan pre/posttreatment significantly increased relative count of limphoblast (p= 0,006; p=0,053; p=0,004), but not spleen weight p=0,277. L-arginine as continuous pretreatment and continuous pre/posttreatment significantly increased phagocytosis (p=0,010; p=0,004), NO production (p=0,037; p=0,010), ROI production (p= 0,016; p=0,030), significantly decreased bacterial count (p=0,004; p=0,006) and significantly increased survival rate (p=0,011; p=0,012). Conclusion: L-arginine supplementation can increase cellular immune response that was consistent with decreasing of bacterial count and increasing of survival rate. It suggests that L-arginine might be useful as immunostimulant against salmonella infection. Key words: L-arginine, lymphocyte proliferation response, phagocytosis, NO, ROI, bacterial count, survival, Salmonella typhimurium Latar belakang: Infeksi Salmonella typhi dapat menekan respon imunitas seluler. Oleh karena itu dalam strategi terapi pada demam tifoid, perlu dipikirkan terapi pendamping dengan imunostimulan. L-arginin, asam amino yang dikenal sebagai prekursor NO dapat meningkatkan respon imun, melalui Growth Hormone maupun poliamin. Tujuan: Untuk mengetahui pengaruh pemberian L-arginin terhadap respon imunitas seluler mencit BALB/c yang diinokulasi Salmonella typhimurium. Metoda: Eksperimental sesungguhnya dengan rancangan the post test only control group design pada mencit BALB/c jantan, sehat, umur 8-10 minggu, berat badan 30-40 gram, diberi makan dan minum adlibitum. Sebanyak 102 ekor mencit setelah diadaptasi 1 minggu dibagi secara acak menjadi 42 ekor untuk pemeriksaan respon imunitas seluler dan 60 ekor untuk pengamatan survival. Empat puluh dua ekor dibagi acak menjadi 7 kelompok perlakuan: A) Kontrol tanpa perlakuan K (-); B) Diinfeksi Salmonella typhimurium (Kontrol+); C) Diberi L-arginin HC1 7 hari tanpa infeksi; D) Diberi Larginin 7 hari pra infeksi kontinyu sebelum infeksi (P2); E) Diberi L-arginin 7 hari pra infeksi selang-seling (P3); F) Diberi L-arginin 7 hari pm infeksi kontinyu dan pasca infeksi kontinyu 3 hari (P4); G) Diberi L-arginin selama 7 hari, sehari pasca infeksi (P5). L-arginin diberikan dalam air minum dengan kadar 1,2% (w/v). Sedang Salmonella typhimurium yang disuntikkan sebanyak 106 intraperitoneal. Kelompok K(+), P2, P3 dan P4 diinfelcsi pada hari ke 8 dan hari ke 11 dibunuh, sedangkan kelompok P5 diinfelcsi hari ke 2 dan dibunuh had ke 8.Kelompok K(-) dan P1 dibunuh pada hari ke 8. Setelah dibunuh dilakukan pemeriksaan respon proliferasi limfosit (dilihat dari bent limpa, jumlah limfosit di limpa dan jumlah relatif limboblas di limpa), kemampuan fagositosis dengan partikel lateks, produksi ROI dengan metode NBT, NO makrofag dengan metode Griess. Hepar diambil dan dilakukan kultur untuk menghitung jumlah kuman yang hidup. Sedangkan survival diamati pada kelompok perlakuan K(+), P2, P3, P4 dan P5 selama 21 hari dengan metode Kaplan Meier. Dosis Salmonella typhimurium yang digunakan untuk pengamatan survival adalah 107 intraperitoneal. Respon imunitas seluler diuji dengan ANOVA dan Kruskal Wallis. Survival ditentukan dengan metoda Kaplan Meier dan diuji dengan Logrank Hasil: Hasil penelitian menunjukkan pemberian L-arginin pm infeksi kontinyu, pm infeksi selang-seling, dan pra/pasca infeksi dapat meningkatkan jumlah limfosit (p=0,037; p=0,037; p=0,010), jumlah relatif limfoblas (p= 0,006; p= 0,053; p= 0,004) secara bermakna, tetapi tidak meningkatkan berat limpa (p=0,277). Pemberian L-arginin pra infeksi kontinyu dan pra/pasca infeksi meningkatkan fagositosis (p=0,010, p=0,004), produksi NO (p=0,037, p=0,010), ROI ( p=0,016, p=0,030), menurunkan hitung kuman (p-0- ,004, p=0,006) dan meningkatkan survival (p= 0,011; p=0,012) secara bermakna. Kesimpulan: Pemberian L-arginin dapat meningkatkan respon imunitas seluler yang sejalan dengan penurunan hitung kuman dan peningkatan survival. L-arginin dapat digunakan sebagai imunostimulan pada infeksi salmonella Kata kunci: L-arginin, respon proliferasi limfosit, fagositosis, NO, ROI, hitung kuman, survival, Salmonella typhimurium.