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"Overtraining meningkatkan IL-1B sistemik akibat mikrotrauma otot sehinggga memengaruhi hipokampus yang penting dalam pembentukan konsolidasi memori spasial. Pemberian H. sabdariffa diharapkan menurunkan IL-1? dan meningkatkan IL-1ra sehingga berpotensi mencegah gangguan konsolidasi memori spasial. Penelitian ini menggunakan metode eksperimental terhadap 20 tikus Wistar jantan Rattus norvegicus, 250-300 gram , terbagi ke dalam 4 kelompok yaitu kontrol C , kontrol H. sabdarifa C-Hib , latihan fisik overtraining OT dan latihan fisik overtraining yang diberi H. sabdarifa OT-Hib . Pemberian ekstrak metanol H. sabdariffa 500 mg/kgBB berpotensi sebagai antiinflamasi melalui peningkatan sitokin antiinflamasi IL-1ra plasma darah secara bermakna sehingga mencegah gangguan fungsi konsolidasi memori spasial tikus overtraining.

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Overtraining lead to increase IL 1 systemically due to muscle mikrotrauma that affect hippocampus which was important in the formation of spatial memory consolidation. Administration H. Sabdariffa is expected to decrease IL 1 and increases IL 1ra thereby potentially preventing impairment of spatial memory consolidation. This research is an experimental study using 20 male Wistar rats Rattus norvegicus, 250 300 g were divided into 4 groups control C , H. sabdarifa control C Hib , physical exercise overtraining OT and physical exercise overtraining by H. sabdarifa OT Hib . Administration of the methanolic extract of H. Sabdariffa 500 mg kg body weight was a potential anti inflammatory by increase anti inflammatory cytokines IL 1ra in blood plasma so that prevent the impairment of spatial memory consolidation in overtraining Wistar rat."

"Insidens insufisiensi adrenal pada pasien renjatan sepsis dilaporkan sekitar 40-65. Sitokin IL-1 dan IL-6 dapat menstimulasi sekresi kortisol sedangkan TNF-? serta MIF berperan dalam menghambat pembentukan kortisol. Penelitian ini bertujuan untuk mengetahui peran IL-1, IL-6, TNF-? dan MIF dalam terjadinya insufisiensi adrenal relatif pada renjatan sepsis.Penelitian eksperimental dilakukan di laboratorium FKH IPB berlangsung selama 6 bulan April-September 2015 . Model anak babi yang dipakai berumur 6-8 minggu dengan berat badan 5-10 kg. Pemilihan sampel dengan consecutive sampling dengan total n = 20. Anak babi diberikan infus endoktoksin dengan dosis 50 ug/kg BB. Sampel darah untuk analisis IL-1, IL-6, TNF-?, MIF, ACTH, kortisol, 17 OHP, DHEA, androstenedion diambil sebelum pemberian endotoksin dan tiap 15 menit hingga terjadi renjatan sepsis, kemudian dilakukan uji synacthen. Pemeriksaan imunohistokimia dilakukan pada kelenjar adrenal, hipofisis, dan hipotalamus.Dari 19 anak babi yang dianalisis mengalami renjatan sepsis dalam waktu 60 menit. Karakteristik sampel tidak berbeda bermakna antara kedua kelompok. Kadar IL-6 pada kelompok IAR dibandingkan dengan kelompok tanpa IAR berbeda bermakna pada menit ke-45 0,65 0,5-4,32 pg/dL vs. 0,54 0,51-0,61 pg/dL , p = 0,008 . Kadar IL-1 antara kelompok IAR dibandingkan kelompok tanpa IAR tidak berbeda bermakna. Kadar TNF-? pada kelompok IAR dibandingkan dengan kelompok tanpa IAR berbeda bermakna pada menit ke-15 1862,5 327,9-4511,14 pg/dL vs. 155,38 24,67-394,10 pg/dL , p = 0,002 dan menit ke-30 4295,76 246,9-5913,37 pg/dL vs. 422,90 101,05-4129,42 pg/dL , p = 0,007 . Kadar MIF kelompok IAR dibandingkan dengan kelompok tanpa IAR berbeda bermakna pada saat renjatan sepsis 25,28 18,45-30,64 ng/dL vs. 11,30 7,1-15,14 ng/dL p = 0,003 . Pemeriksaan imunohistokimia hanya pada hipotalamus yang menunjukkan pewarnaan terhadap IL-1, IL-6, TNF-? dan MIF pada kelompok dengan IAR. Pada renjatan sepsis dan insufisiensi adrenal relatif kadar TNF-? meningkat pada menit-menit awal, kemudian kadar IL-6 meningkat kemudian serta terakhir kadar MIF meningkat pada saat renjatan sepsis. Kadar IL-1 tidak terdapat perbedaan antara kedua kelompok. Kata kunci: IL-1, IL-6, insufisiensi adrenal relatif, MIF, renjatan sepsis, TNF-?

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Incidence of adrenal insufficiency in septic shock ranged between 40 ndash 65 . The mechanism of relative adrenal insufficiency in septic shock is caused by inflammatory mediators. This study aimed to identify the role of IL 6, IL 1 in stimulating ACTH and cortisol release, and the role of TNF and MIF in inhibiting the level of ACTH and cortisol in septic shock with relative adrenal insufficiency RAI in order to develop guidelines for relative adrenal insufficiency marker.Experimental study was conducted in Veterinary Faculty, Bogor Agricultural Institute for 6 months Apri ndash September 2015 . Piglet models Sus scrofa aged 6 ndash 8 weeks weighing 5 ndash 10 kg. Consecutive sampling was used with total 20 piglets. Piglet models were given 50 ug kg endotoxin infusion Escherichia coli O111 B4 Sigma chemical, St. Louis, MO, USA . Blood sample for analysis of IL 1, IL 6, TNF , MIF, ACTH, cortisol, 17 OHP, DHEA, androstenedione was collected before endotoxin administration and every 15 minutes until septic shock occurred. Piglet models were monitored using PiCCO monitor. Stimulation test was then performed using synthetic corticotropin Synacthen and blood sample was collected again along with immunohistochemistry examination of the adrenal, pituitary and hypothalamus glands.From 19 study subjects analized, all subject had septic shock in 60 minutes. Study subject characteristics in each group were similar. The level of IL 6 at 45 minutes had a significant different compared to the group without RAI 0.65 0.5 ndash 4.32 pg dL vs. 0.54 0.51 ndash 0.61 pg dL , p 0.008 . The level of IL 1 during septic shock were not significantly different between both groups. The level of TNF in RAI group had significant different compared to the group without RAI at 15 minutes 1862.5 327.9 ndash 4511.14 pg dL vs. 155.38 24.67 ndash 394.10 pg dL , p 0.002 and at 30 minutes 4295.76 246.9 ndash 5913.37 pg dL vs. 422.90 101.05 ndash 4129.42 pg dL , p 0.007 The level of MIF in group with RAI during septic shock had a significant different compared to the group without RAI t 25.28 18.45 ndash 30.64 ng dL vs. 11.30 7.1 ndash 15.14 ng dL , p 0.003 . Immunohisto chemistry staining of IL 1, IL 6, TNF , and MIF was observed only in the hypothalamus glands of the RAI group. In septic shock and relative adrenal insufficiency, TNF increased in earlier minutes, then IL 6 increased and later MIF increased in septic shock condition. IL 1 level had no difference increment for both group.Keywords IL 1, IL 6, MIF, relative adrenal insufficiency, septic shock, TNF"

"Pengaruh Ekstrak Daun Jintan (Plectranthus amboinicus) terhadap Kadar BUN dan Kreatinin serta Respon Seluler Faktor Proinflamasi TNF-α dan IL-1β Pasien Gout Artritis. Pengobatan gout artritis (GA) saat ini dikembangkan berbasis anti sitokin yaitu blokade kemokin, penghambatan pelepasan IL-1β dan TNF-α. Tujuan penelitian ini adalah mengembangkan pengobatan berbasis anti sitokin dengan menggunakan ekstrak daun jintan yang diaplikasikan pada penderita GA. Penelitian ini adalah penelitian eksperimental semu dengan desain penelitian randomized pretest-posttest control group design. Pengambilan sampel pasien GA di Instalasi Rawat Jalan Poli Penyakit Dalam RSU Haji Surabaya yang memenuhi kriteria inklusi dan eksklusi. Jumlah sampel sebanyak 30 responden terbagi menjadi kelompok perlakuan dan kelompok kontrol masing-masing 15 responden. Pada kelompok perlakuan diminta untuk minum obat dari Rumah Sakit ditambah dengan kapsul ekstrak daun jintan selama 7 hari, dengan dilakukan pengamatan keradangan sendi. Pada kelompok kontrol minum obat dari Rumah Sakit dan dilakukan pengamatan keradangan sendi. Sampel darah diambil sebelum dan sesudah perlakuan untuk mengukur kadar BUN, kreatinin, konsentrasi TNF-α dan IL-1β. Terdapat penurunan kadar BUN dan kreatinin pada kelompok kontrol namun tidak signifikan yaitu menurun sebesar 3% dan 8%. Sementara pada kelompok perlakuan terdapat peningkatan kadar BUN dan Kreatinin namun juga tidak signifikan sebesar 3% dan 7%. Terdapat penurunan konsentrasi TNF-α pada kelompok kontrol sebesar 9% dan kelompok perlakuan sebesar 22%, keduanya tidak signifikan. Sementara konsentrasi IL-1β terjadi peningkatan pada kelompok kontrol sebesar 18%, sementara pada kelompok perlakuan terjadi penurunan sebesar 3%, keduanya tidak signifikan.

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The purpose of this research was to develop anti-cytokine-based treatment using extract of Plectranthus amboinicus applied to gout arthritis (GA) patients. The research was quasi experimental, with a pretest-posttest randomized control group design. The samples were GA patients in the Outpatient Installation of Internal Medicine in General Hospital Haji, Surabaya. The sample was comprised of 30 respondents. The respondents were divided into a treatment group and a control group. The treatment group was asked to take medicine from the hospital, coupled with P. amboinicus extract capsules, for 7 days, during which time patients? joint inflammation was observed. The control group was provided with only medication from the hospital, and their joint inflammation was likewise observed. Blood samples were taken before and after treatment, to measure the levels of blood urea nitrogen (BUN) and creatinine, as well as the concentrations of TNF-α and IL-1β. There was a decrease in BUN and creatinine levels in the control group, but it was not significant, decreasing by 3% and 8%, respectively. The treatment group also showed elevated levels of BUN and creatinine, which also was not significant at 3% and 7%, respectively. There was a decrease in the concentration of TNF-α in the control group by 9% and 22%. The concentration of IL-1β in the control group increased by 18%, whereas,in the treatment group, it decreased by 3%; however, the decreases in both groups were not significant."

"The purpose of this research was to develop anti-cytokine-based treatment using extract of Plectranthus amboinicusapplied to gout arthritis (GA) patients. The research was quasi experimental, with a pretest-posttest randomized controlgroup design. The samples were GA patients in the Outpatient Installation of Internal Medicine in General HospitalHaji, Surabaya. The sample was comprised of 30 respondents. The respondents were divided into a treatment group anda control group. The treatment group was asked to take medicine from the hospital, coupled with P. amboinicus extractcapsules, for 7 days, during which time patients’ joint inflammation was observed. The control group was provided withonly medication from the hospital, and their joint inflammation was likewise observed. Blood samples were takenbefore and after treatment, to measure the levels of blood urea nitrogen (BUN) and creatinine, as well as theconcentrations of TNF-α and IL-1β. There was a decrease in BUN and creatinine levels in the control group, but it wasnot significant, decreasing by 3% and 8%, respectively. The treatment group also showed elevated levels of BUN andcreatinine, which also was not significant at 3% and 7%, respectively. There was a decrease in the concentration ofTNF-α in the control group by 9% and 22%. The concentration of IL-1β in the control group increased by 18%,whereas,in the treatment group, it decreased by 3%; however, the decreases in both groups were not significant.Pengaruh Ekstrak Daun Jintan (Plectranthus amboinicus) terhadap Kadar BUN dan Kreatinin serta ResponSeluler Faktor Proinflamasi TNF-α dan IL-1β Pasien Gout Artritis. Pengobatan gout artritis (GA) saat inidikembangkan berbasis anti sitokin yaitu blokade kemokin, penghambatan pelepasan IL-1β dan TNF-α. Tujuanpenelitian ini adalah mengembangkan pengobatan berbasis anti sitokin dengan menggunakan ekstrak daun jintan yangdiaplikasikan pada penderita GA. Penelitian ini adalah penelitian eksperimental semu dengan desain penelitianrandomized pretest-posttest control group design. Pengambilan sampel pasien GA di Instalasi Rawat Jalan PoliPenyakit Dalam RSU Haji Surabaya yang memenuhi kriteria inklusi dan eksklusi. Jumlah sampel sebanyak 30responden terbagi menjadi kelompok perlakuan dan kelompok kontrol masing-masing 15 responden. Pada kelompokperlakuan diminta untuk minum obat dari Rumah Sakit ditambah dengan kapsul ekstrak daun jintan selama 7 hari,dengan dilakukan pengamatan keradangan sendi. Pada kelompok kontrol minum obat dari Rumah Sakit dan dilakukanpengamatan keradangan sendi. Sampel darah diambil sebelum dan sesudah perlakuan untuk mengukur kadar BUN,kreatinin, konsentrasi TNF-α dan IL-1β. Terdapat penurunan kadar BUN dan kreatinin pada kelompok kontrol namuntidak signifikan yaitu menurun sebesar 3% dan 8%. Sementara pada kelompok perlakuan terdapat peningkatan kadarBUN dan Kreatinin namun juga tidak signifikan sebesar 3% dan 7%. Terdapat penurunan konsentrasi TNF-α padakelompok kontrol sebesar 9% dan kelompok perlakuan sebesar 22%, keduanya tidak signifikan. Sementara konsentrasiIL-1β terjadi peningkatan pada kelompok kontrol sebesar 18%, sementara pada kelompok perlakuan terjadi penurunansebesar 3%, keduanya tidak signifikan."

"Tujuan umum: Mengetahui profil keamanan dan efek getah J. curcas terhadap jaringan gigi dan periapeks dalam persiapan untuk memanfaatkan pemakaian bahan alami getah J. curcas pada radang pulpa. Tujuan khusus (1) Mengetahui kandungan golongan senyawa getah J. curcas. (2) Mengetahui sitotoksisitas getah J. curcas. (3) Mengetahui toksisitas akut pemberian secara oral dosis tunggal getah J. curcas pada hewan percobaan. (4) Mengetahui aktivitas hemolisis getah J. curcas pada darah manusia secara in vitro. (5) Mengetahui sifat mutagenisitas getah J. curcas. (6) Mengetahui efek getah J. curcas terhadap pembebasan interleukin-1β oleh sel makrofag. (7) Mengetahui efek getah J. curcas terhadap pembebasan kolagenase pada set fibroblast. (8) Mengetahui efek histopatologik getah J. curcas terhadap pulpa dan jaringan periapeks gigi pada hewan percobaan. (9) Mengetahui efek getah J. curcas terhadap kekerasan macro jaringan keras gigi manusia secara in vitro. (10) Mengetahui efek getah J. curcas terhadap jaringan keras gigi manusia dalam hal kelarutan unsur kalsium dan fosfat secara in vitro. Metode penelitian: Disain penelitian eksperimental dan eksplorasi. Penelitian dibagi atas (1) skrining fitokimia, (2) tahap 1 dan (3) tahap 2 evaluasi biologik getah J. curcas. Untuk standardisasi getah J. curcas diambil dari satu petak tanaman dalam satu musim, kemudian diukur pH, volume basah, diliofilisasi, diukur berat kering, dan disimpan pada -20°C sebagai sampel. (1). Skrining fitokimia getah J. curcas. Analisis kualitatif golongan senyawa diidentifikasi dari ekstrak eter, etil asetat, dan air. (2). Uji toksisitas 1. Uji sitotoksisitas. (1) Metoga agar overlay. Getah J. curcas dan kontrol diserap oleh cakram selulosa, kemudian diletakkan di atas permukaan agar yang menutupi selapis sel Fib L929 yang telah diwarna neutral red. Evaluasi berdasar luas zona dekolorisasi dan zona lisis yang terbentuk setelah 24 jam. (2) Assay MTT. Getah J. curcas dalam medium diberikan pada kultur set Fib L929 cell line dan sel primer fibroblast gingiva manusia yang tumbuh dalam mikroplat 96-sumur. Setelah 1-4 hari, dilakukan assay MTT. Evaluasi berdasar perbandingan nilai OD kontrol dan perlakuan. 2. Uji toksisitas akut. Mencit diberi getah J. curcas secara intragastrik sebanyak 1 kali. Dihitung LD5O berdasar jumlah mencit yang mati. Dibandingkan antara kelompok perlakuan dan kontrol dalam hal tanda toksisitas, berat badan selama 2 minggu, pemeriksaan makroskopik dan mikroskopik organ tubuh. 3. Uji hemolisis. Darah dicampur dengan berbagai konsentrasi getah J. curcas. Evaluasi berdasar pembebasan hemoglobin, dibandingkan OD kelompok perlakuan dengan kontrol positif air, dan kontrol negatif salin. 4. Uji mutagenisitas. Getah J. curcas dikultur dengan bakteri S. typhi dan E. coil mutan. Evaluasi berdasar penghitungan koloni reversi bakteri, dibandingkan kelompok perlakuan, kontrol positif dan kontrol negatif. (3) Efek getah J. curcas terhadap makrofag dan fibroblast 1. Efek getah J. curcas terhadap pembebasan IL-1β. Lima dosis getah J. curcas dimasukkan ke dalam kultur makrofag peritoneum mencit BALB/c, secara bersamaan, sebelum, atau sesudah pemberian LPS. Setelah 1 dan 2 hari, IL-1β dalam supernatan diukur secara ELISA dengan Quantikine IL-1β for mouse kit. 2. Efek getah J. curcas terhadap pembebasan kolagenase oleh fibroblast. Empat dosis getah J. curcas dan IL-1β dimasukkan dalam kultur sel primer fibroblast gingiva manusia. Setelah 1-4 hari kolagenase dalam supematan diukur dengan assay kolagenase. Hasil degradasi kolagen dipisahkan dengan SDS-PAGE. Pita 3/4 αA diukur dengan program komputer Adobe Photo. (4) Efek histopatologik getah J. curcas pada jaringan pulpa dan periapeks. Getah J. curcas dimasukkan ke dalam kavitas gigi monyet. Setelah 3 hari, gigi diproses untuk pembuatan sediaan histologik. Evaluasi berdasar perbandingan pemeriksaan keadaan mikroskopik jaringan pulpa dan peripeks dalam hal inflamasi dan nekrosis, antara kelompok kontrol dan perlakuan. (5) Efek getah J. curcas terhadap jaringan keras gigi. 1. Efek getah J. curcas terhadap kekerasan mikro dentin dan email. Mahkota gigi premolar dibelah 4 longitudinal, lalu ditanam di dalam akrilik dengan 1 permukan tidak tertutup akrilik. Setelah direndam dalam 3 konsentrasi getah J. curcas, permukaan dentin dan email diberi indentasi oleh intan Knoop. Evaluasi berdasar perbandingan KHN kelompok kontrol dan perlakuan. 2. Efek getah J. curcas terhadap kelarutan kalsium dan fosfat. Mahkota gigi premolar utuh dibelah 4 secara longitudinal, lalu direndam dalam 3 konsentrasi getah J. curcas. Setelah 1-3 hari, kalsium dan fosfat yang larut dalam rendaman diukur berturut-turut dengan alat atomic absorption spectrophotometer (AAS) dan spektrofotometer (metoda asam askorbat). Hasil penelitian pH getah J. curcas rata-rata 3,49 ± 0,09 dan perbandingan berat kering/volume basah 15,12 ± 0,31%. (1) Skrining fitokimia: getah J. curcas mengandung golongan senyawa sterol, aglikon flavon, tanin, senyawa pereduksi, glikosida steroid, poliose, dan saponin. (2) Uji toksisitas 1.(1) Sitotoksisitas getah J. curcas pada metoda agar overlay ditemukan zona dekolorisasi indeks 2 dari 5 indeks zona. Tak ada lisis sel, bentuk sel masih jelas. (2) Assay MTT: pads getah J. curcas kadar 0,25% terhadap Fib L929 dan kadar 0,12% terhadap fibroblast gingiva, sel nekrosis. 2.(1) LD50 > 5 g/kg BB, sehingga getah J. curcas dapat diklasifikasi dalam toksik ringan. (2) Tidak ada perbedaan berat badan. (3) Tidak ada perbedaan makroskopik dan mikroskopik organ tubuh yang diperiksa. (4) Terjadi inaktivitas pada hari 1 pada kelompok perlakuan, selanjutnya tidak ada perbedaan. 3. Aktivitas hemolisis getah J. curcas 15% adalah 6,5% dibanding air. Tidak ada hemolisis pada konsentrasi getah J. curcas yang lebih rendah. 4. Tidak ada aktivitas mutagenisitas getah J. curcas. (3) Efek getah J. curcas terhadap makrofag dan fibroblast 1. (1) LPS meningkatkan pembebasan 1L-1β oleh makrofag. (2) Pemberian getah J. curcas menghambat pembebasan 1L-1β oleh makrofag. 2. (1) Makin lama waktu kultur, produksi kolagenase makin banyak. (2) Getah J. curcas menurunkan pembebasan kolagenase oleh fibroblast. (4) Efek histopatologik getah J. curcas terhadap jaringan pulpa dan periapeks (1) Inflamasi dan nekrosis terj adi pads daerah yang terbatas dekat dengan daerah yang kontak dengan getah J. curcas. Di bawahnya terdapat jaringan pulpa normal. (2) Tingkat inflamasi pulpa kelompok perlakuan tidak lebih parah dari kelompok kontrol. (3) Tidak ada radang periapeks pads kelompok kontrol dan perlakuan. (5) Efek getah J. curcas terhadap jaringan keras gigi. 1. Efek getah J. curcas terhadap kekerasan mikro dentin dan email. (1) Kekerasan mikro dentin tidak berbeda bermakna pada 1 dan 2 hari perendaman getah J. curcas antara kelompok kontrol dan perlakuan. Namur lebih kecil setelah 3 hari pada konsentrasi getah 15%. (2) Kekerasan mikro email tidak berbeda antara kelompok kontrol dan perlakuan pada 1 dan 3 hari, Namun lebih kecil setelah 2 hari pada konsentrasi getah J. curcas 15%. 2. Kadar kalsium dan fosfat yang larut meningkat sesuai dengan kenaikan konsentrasi getah J. curcas. Namun lama perendaman tidak berpengaruh secara bermakna terhadap kelarutan kalsium. Kesimpulan (1) Getah J. curcas mengandung sterol, aglikon flavon, tanin, senyawa pereduksi, glikosida steroid, poliose, dan saponin. (2) Tahap 1 evaluasi biologik: getah J. curcas relatif aman pada hewan percobaan berdasar LD50>5 g/kg BB sehingga termasuk dalam klasifkasi toksik ringan; hemolisis 6,5% dibanding air; tidak mutagen; dan sitotoksik dengan nekrosis koagulasi. (3) Uji tahap 2: getah J. curcas cukup efektif dalam menanggulangi pulpalgia, berdasar nekrosis pulpa terbatas, tidak ada kelainan periapeks; kekerasan mikro email dan dentin tidak turun pada 1 hari; menghambat pembebasan IL-1β dan kolagenase. Namun getah melarutkan kalsium dan fosfat. Kesimpulan penelitian: penelitian dapat dilanjutkan ke tahap uji klinik atau tahap 3.

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Biological Study on the Effects of Jatropha Curcas (Euphorbiaceae) Latex on Dental and Periapical TissuesObjective: The objective of this study was to evaluate the safety level and the effects of J. curcas latex on dental and periapical tissues. The aims in details were (1) to identify the main classes of chemical constituent in J. curcas latex; (2) to evaluate the cytotoxicity of J. curcas latex; (3) to determine the acute toxicity of J. curcas latex after single oral administration on mice; (4) to assess hemolytic activity of J. curcas latex; (5) to evaluate mutagenic activity of J. curcas latex; (6) to evaluate the effect on J. curcas latex of IL-1 il release from macrophages; (7) to evaluate the effect of J. curcas latex on collagenase release from fibroblasts; (8) to assess the histopathological effects of J. curcas latex on monkey dental pulp and periapical tissues; (9) to determine the effects of J. curcas latex to dentin and enamel micro-hardness; (10) to assess the effects of J. curcas latex on dissolving calcium and phosphate. Methods: Research design was experimental and explorative. To standardize the sample, J. curcas latex was collected from Balittro, Bogor in 1997, then the pH and wet volume were measured, the latex was lyophilized, dry weight was measured, and latex was stored at-20°C as sample. Biological evaluation was grouped into (1) phytochemical sreening, (2) toxicity test, (3) effects of J.curcas latex on cell, (4) effects of J.curcas latex on dental pulp and periapical tissues, and (5) effects of J.curcas latex on dental hard tissues, (1). Phytochemical screening: the main classes of chemical constituents of J. curcas latex were analyzed qualitatively from ether, ethyl acetate, and water extracts. (2). Toxicity test 1. Cytotoxicity test. (1) Agar overlay technique. J. curcas latex was imbibed in cellulose discs and put on the surface of agar overlaying a neutral red stained Fib L929 cell monolayer. Evaluation was judged on zone index and lysis index after 24 hours incubation. (2) MT assay. J. curcas latex was added to human gingival fibroblasts and Fib L929 cell culture in 96-well micro-plates. After 1-4 days of incubation, MTT assay was performed. Evaluation was based on comparing the OD values of control and test groups. 2. Acute toxicity. A single dose of J. curcas latex was given to male and female mice, intragastrically. LD50 was determined based on mortality rate. Assessment was also performed on 2 weeks observations of body weight, macroscopic and microscopic examinations of several organs. 3. Hemolysis test. Blood was mixed with several concentrations of J. curcas latex. The result was the extent of hemolysis expressed based on the absorbance of the test samples, negative and positive controls. 4. Mutagenicity test. L curcas latex was added to the S. ryphi and E. coil mutans culture. Assessment was based on bacterial revertant colonies, compare to positive and negative controls. (3) Effects of J.curcas latex on macrophages and fibroblasts 1. Effects of .T. curcas latex on the release of IL-1 β from macrophages. Five doses of J. curcas latex from 75-1200 μg/ml were added into the culture of BALB/c mice peritoneal macrophages, along with, after, or before addition of LPS. Following 1-3 days of incubation, IL-1P presence in supernatant was measured by ELISA using Quantikine ]L-1P for mouse kit. 2. Effects of J. curcas latex on the release of collagenase. Four doses of J. curcas latex from 37.5-300 µg/ml were added to human gingival fibroblasts cell culture. After 1-4 days of incubation, collagenase in the supernatant was assayed with collagen. The degradation products were then separated by SDS-PAGE and the density of 3/4 αA bands was measured semi quantitatively by Adobe Photo computer program. (4) Effects of J.curcas latex on dental pulp and periapical tissues. The latex of J. curcas was brought in contact with dental pulp and sealed. Assessment was based on the presence of inflammation and necrosis in dental pulp and periapical tissues, histopathologically. (5) Effects of J.curcas latex on dental hard tissues 1. Effects of J. curcas latex on dentin and enamel micro-hardness. Intact premolar crowns were cut longitudinally into 4 fragments, followed by embedding of each fragment in acrylats leaving 1 free surface. The fragments were then soaked in 3 concentrations of J. curcas latex from 3.7-15% for 1-3 days. The dentin and enamel micro-hardness were assessed by Knoop hardness measurement. 2. Effects of J. curcas latex on dissolved calcium and phosphate. Intact premolar crowns were cut longitudinally into 4 fragments, followed by soaking the fragments in 3 concentration of J. curcas latex from 3.7-15% for 1-3 days. The dissolved calcium and phosphate were measured according to atomic absorption spectrophotometer and spectrophotometer (ascorbic acid method), respectively. Results: The mean ± SD of J. curcas latex pH was 3.49 ± 0.09. The dry weight/wet volume was 15.12 ± 0.31%. (1). Phytochemical screening: sterols, flavone aglycones, tannins, reducing compounds, sterol glycosides, poliose, and saponins were identified in J. curcas latex. (2) Toxicity test 1. (1) Agar overlay technique. 2-5 mm decoloration zones were observed, indicating that J. curcas latex was cytotoxic. No lysis of cells was observed within the decolorized zone. (2) MTT assay. At 2.5 mg/ml J. curcas latex no living Fib L929 cells were observed, while the same result was shown at 1.2 mg/ml J. curcas latex on human gingival fibroblasts. 2. LD50 was more than 5 g/kg BW, hence dry J. curcas latex may be classified into mildly toxic substance. No significant body weight difference was observed. Macroscopic and microscopic examination on several organs showed no differences between test and control groups. 3. 6,5% hemolytic activity of 15% J. curcas latex compared to water was observed, while no hemolisis was observed with lower concentrations of latex. 4. No mutagenic ativity was observed with J. curcas latex. (3) Effects of J.curcas latex on macrophages and fibroblasts 1. (1) LPS increased the release of IL-1β. (2) J. curcas latex inhibited the release of IL-lβ from macrophages. 2. (1) The longer the duration of incubation, the more collagenase was released. (2) J. curcas latex decreased collagenase release by human gingival fibroblast. (4) Effects of I. curcas latex on dental pulp and periapical tissues. Inflammation and necrosis were observed in a limited area, which was in direct contat with J. curcas latex, underneath was normal pulp. Inflammation in the pulp of test group was not greater than in the control group. No inflammation or necrosis in periapical tissues was observed in all groups. (5) Effects of J. curcas latex on dental hard tissues 1. (1) The micro-hardness of dentin was not lowered after 1 and 2 days treatment, but lower after 3 days at 15% J. curcas latex. (2) The enamel microhardness was not decreased after 1 and 3 days immersion in J. curcas latex, but decreased after 2 days at 15% J. curcas latex. 2. The calcium and phosphate release were increased in accordance to the concentration of J. curcas latex. The duration of treatment did not influence the release of calcium, while it influenced the release of phosphate. Conclusions (1) J. curcas latex contains sterols, flavone aglycones, tannins, reducing compounds, sterol glycosides, poliose, and saponins. (2) Level 1 biological evaluation: J. curcas latex is relatively safe in animals based on LD50>5 g/kg BW, 6,5% hemolysis compared to water, not mutagenic, but cytotoxic with coagulative necrosis. (3) Level 2 biological evaluation: J. curcas latex seems to be effective in relieving pulpal pain. It caused coagulative necrosis in pulp, which was in direct contact with J. curcas latex while the tissue underneath was normal. It did not cause inflammation of periapical tissues, and did not lower the dentin and enamel micro-hardness after 1 day of exposure, but it lowered the microhardness after 3 days. It inhibited IL-1β and collagenase release. It dissolved dental calcium and phosphate."