Periodontite é uma doença inflamatória crônica comum caracterizada pela perda de osso alveolar. O alto teor de polifenóis na casca do cacau (Theobroma cacao L) tem o potencial de influenciar o metabolismo ósseo e contribuir para a inibição da reabsorção óssea. O objetivo deste estudo foi analisar o potencial anti-osteoclastogênico da casca do cacau (Theobroma cacao L.) em um estudo in silico e in vivo. Uma análise do potencial anti-osteoclastogênico dos compostos bioativos do T. cacao foi realizada por simulações de docking molecular. Trinta ratos Wistar machos (Rattus novergicus) foram aleatoriamente selecionados em grupos controle negativo (gel placebo), controle positivo (gel de doxiciclina a 2%) e grupos de tratamento (gel com extrato de casca de cacau a 10%), e as medições foram feitas nos dias 7 e 14. Ratos Wistar foram induzidos com 0,05 ml de P. gingivalis a uma concentração de 2x10^9 UFC/ml de forma intrasulcular no molar maxilar para atingir a periodontite. O número de osteoclastos foi observado por coloração de hematoxilina e eosina, o nível de TNF-α foi observado por ensaio imunoenzimático, e a expressão de RANKL foi observada por coloração imunohistoquímica. Os dados foram analisados usando ANOVA de uma via para examinar a diferença entre os grupos. O estudo in silico mostrou que a catequina, epicatequina, quercetina e procyanidina B2 tinham uma forte afinidade de ligação para TNF-α e RANKL. A administração de 10% de CPH reduziu o número de osteoclastos (p<0,05), o nível de TNF-α nos dias 7 e 14 (p<0,05) e a expressão de RANKL no dia 7 (p<0,05) em ratos experimentais com periodontite. A administração de 10% de CPH inibiu a osteoclastogênese nos ratos com periodontite experimental
Autoria
person Yani Corvianindya Rahayu
schoolDoctoral Study Program in Dental Science, Faculty of Dental Medicine, Universitas Airlangga, Surabaya, Indonesia Universitas AirlanggaIndonesiaSurabaya, IndonesiaDoctoral Study Program in Dental Science, Faculty of Dental Medicine, Universitas Airlangga, Surabaya, Indonesia schoolDepartment of Oral Biology, Faculty of Dentistry, Universitas Jember, IndonesiaUniversitas JemberIndonesiaIndonesiaDepartment of Oral Biology, Faculty of Dentistry, Universitas Jember, Indonesia
person Ernie Maduratna Setiawatie
person Retno Pudji Rahayu
person S Siswandono
person Retno Indrawati
person Hendrik Setia Budi
person Hari Basuki Notobroto
schoolDepartment of Epidemiology, Biostatistics and Population Studies, and Health Promotion, Faculty of Public Health, Universitas Airlangga, Surabaya, Indonesia Universitas AirlanggaIndonesiaSurabaya, IndonesiaDepartment of Epidemiology, Biostatistics and Population Studies, and Health Promotion, Faculty of Public Health, Universitas Airlangga, Surabaya, Indonesia
person Rahmah A. Alwasilah Darojah
Correspondence: Yani Corvianindya Rahayu, Department of Oral Biology, Faculty of Dentistry, Universitas Jember, jl. Kalimantan no.37 Jember, Indonesia, +62 8113536391.
SCIMAGO INSTITUTIONS RANKINGS
Doctoral Study Program in Dental Science, Faculty of Dental Medicine, Universitas Airlangga, Surabaya, Indonesia Universitas AirlanggaIndonesiaSurabaya, IndonesiaDoctoral Study Program in Dental Science, Faculty of Dental Medicine, Universitas Airlangga, Surabaya, Indonesia
Department of Oral Biology, Faculty of Dentistry, Universitas Jember, IndonesiaUniversitas JemberIndonesiaIndonesiaDepartment of Oral Biology, Faculty of Dentistry, Universitas Jember, Indonesia
Department of Periodontology, Faculty of Dental Medicine, Universitas Airlangga, Surabaya, Indonesia Universitas AirlanggaIndonesiaSurabaya, IndonesiaDepartment of Periodontology, Faculty of Dental Medicine, Universitas Airlangga, Surabaya, Indonesia
Department of Oral Pathology and Maxillofacial, Faculty of Dental Medicine, Universitas Airlangga, Surabaya, IndonesiaUniversitas AirlanggaIndonesiaSurabaya, IndonesiaDepartment of Oral Pathology and Maxillofacial, Faculty of Dental Medicine, Universitas Airlangga, Surabaya, Indonesia
Department of Pharmacology, Faculty of Pharmacy, Universitas Airlangga, Surabaya, IndonesiaUniversitas AirlanggaIndonesiaSurabaya, IndonesiaDepartment of Pharmacology, Faculty of Pharmacy, Universitas Airlangga, Surabaya, Indonesia
Department of Oral Biology, Faculty of Dental Medicine, Universitas Airlangga, Surabaya, IndonesiaUniversitas AirlanggaIndonesiaSurabaya, IndonesiaDepartment of Oral Biology, Faculty of Dental Medicine, Universitas Airlangga, Surabaya, Indonesia
Department of Epidemiology, Biostatistics and Population Studies, and Health Promotion, Faculty of Public Health, Universitas Airlangga, Surabaya, Indonesia Universitas AirlanggaIndonesiaSurabaya, IndonesiaDepartment of Epidemiology, Biostatistics and Population Studies, and Health Promotion, Faculty of Public Health, Universitas Airlangga, Surabaya, Indonesia
Figuras | Tabelas
imageFigure. 1 Histological features of alveolar bone resorption in the maxillary first molar of a Wistar Rat periodontitis model and measurement area in the buccal periosteum (BP). A) hematoxyllin eosin staining B) immunohistochemistry staining open_in_new

imageFigure 2 Three-dimensional (3D) visualization of docking results of four selected ligands with TNFR (A) Diclofenac, (B) Catechin, (C) (-)-Epicatechin (D) Procyanidin B2, (E) Quercetin. open_in_new

imageFigure. 3 Three-dimensional (3D) visualization of docking results of four selected ligands with RANKL (A) Diclofenac, (B) Catechin, (C) (-)- Epicatechin (D) Procyanidin B2, (E) Quercetin. open_in_new

imageFigure. 4 Histological features of alveolar bone’s periodontitis rats osteoclast in hematoxillin eosin staining at 400x magnification. a) negative control group (placebo CMC-Na gel) day 7; b) positive control group (doxycycline gel) day 7; c) treatment group (CPH 10% gel) day 7; d) negative control group (placebo CMC-Na gel) day 14; e) positive control group (doxycycline gel) day 14; f) treatment group (CPH 10% gel) day 14. Black arrows indicated osteoclast cells. open_in_new

imageFigure. 5 Histological features of RANKL immunohistochemistry at 400x magnification on alveolar bone of periodontitis rats. a) negative control group (placebo CMC-Na gel) day 7; b) positive control group (doxycycline gel) day 7; c) treatment group (CPH 10% gel) day 7; d) negative control group (placebo CMC-Na gel) day 14; e) positive control group (doxycycline gel) day 14; f) treatment group (CPH 10% gel) day 14. Black arrows indicated RANKL-expressing osteoblast cells. open_in_new

table_chartTable 1
Molecular docking simulation and interaction of selected phytochemicals of Theobroma cacao with TNFR
| Protein target | Compounds (ligands) | Binding affinity [kcal/mol] | Hydrogen bonds | van der Waals interaction |
|---|---|---|---|---|
| TNFR | Diclofenac (3033) | -6.2 | G1n149, Val146 | Ala222, Leu102, Leu147,Ser145, Val148 |
| Catechin (9064) | -6.5 | Leu102, Trp226 | Gln149, Gln22, Gly70, His224, Ile103, Leu147, Val152, Val71 | |
| (-)-Epicatechin (72276) | -6.1 | Leu102 | Ala222, Ala225, Gln149, His224, Leu101, Ser145 | |
| Quercetin (5280343) | -6.5 | His224, Leu147 | Gln227, Gly70, Ile103, Leu102, Ser145, Trp226, Val148, Val152, Val71 | |
| Procyanidin B2 (122738) | -5.4 | Arg223, His224, Trp226 | Ala222, Gln149, Leu102, Leu147, Ser135, Val152, Val71 |
table_chartTable 2
Molecular docking simulation and interaction of selected phytochemicalsn of Theobroma cacao with RANKL
| Protein target | Compounds (ligands) | Binding affinity [kcal/mol] | Hydrogen bonds | van der Waals interaction |
|---|---|---|---|---|
| RANKL | Raloxifene (5035) | -7.6 | Glu287, Tyr61 | Arg284, Ile249, Leu255, Leu60, Met256, Pro64, Ser63, Tyr48 |
| Catechin (9064) | -6.7 | Ser63, Thr254 | Arg284, Glu58, Ile249, Leu255, Leu60, Lys248, Lys282, Ser246, Tyr61 | |
| (-)-Epicatechin (72276) | -7.0 | Cys59, Ser63, Thr254 | Glu287, Glu58, Ile249, Leu255, Lys244, Lys248, Ser246, Tyr48, Tyr61 | |
| Quercetin (5280343) | -6.8 | Glu58, Tyr61 | Arg284, Cys59, His253, Ile249, Leu255, Lys244, Phe281, Thr254 | |
| Procyanidin B2 (122738) | -6.9 | Tyr61 | Ile249, Leu255, Leu60, Lys244, Lys257, Lys282, Met256, Pro64, Ser63m Thr254 |
table_chartTable 3
The level of TNF-α in gingival crevicular fluids of periodontitis rat
| Day | TNF-α (mean ± SD) | p value | ||
|---|---|---|---|---|
| NC | PC | CPH | ||
| 7 | 146.72 ± 5.91* | 96.15 ± 5.39 | 99.44 ± 3.32 | 0.001 |
| 14 | 114.72 ± 4.72* | 75.59 ± 4.86 | 79.52 ± 3.51 | 0.000 |
-
All data were collected in 3 groups (n=5). NC= Negative control group administering CMC-Na gel on day 7 and 14; PC= Positive control group administering doxycycline gel on day 7 and 14; CPH= Treatment group administering cocoa pod husk extract gel on day 7 and 14. *Significant p values (p ≤ 0.05).
table_chartTable 4
The number of osteoclast on alveolar bone of periodontitis rats
| Day | Osteoclast (mean ± SD) | p value | ||
|---|---|---|---|---|
| NC | PC | CPH | ||
| 7 | 6.25 ± 0.95* | 3.50 ± 1.29* | 3.00 ± 0.81* | 0.004 |
| 14 | 3.50 ± 0.57* | 1.75 ± 0.95 | 1.75 ± 0.95 | 0.047 |
-
All data were collected in 3 groups (n=5). NC= Negative control group administering CMC-Na gel on day 7 and 14; PC= Positive control group administering doxycycline gel on day 7 and 14; CPH= Treatment group administering cocoa pod husk extract gel on day 7 and 14. *Significant p values (p ≤ 0.05).
table_chartTable 5
The expression of RANKL on alveolar bone of periodontitis rats
| Day | RANKL (mean ± SD) | p value NC | ||
|---|---|---|---|---|
| NC | PC | CPH | ||
| 7 | 18.75 ± 2.62* | 12.00 ± 0.81* | 7 | 18.75 ± 2.62* |
| 14 | 15.50 ± 1.29* | 9.00 ± 0.81* | 14 | 15.50 ± 1.29* |
-
All data were collected in 3 groups (n=5). NC= Negative control group administering CMC-Na gel on day 7 and 14; PC= Positive control group administering doxycycline gel on day 7 and 14; CPH= Treatment group administering cocoa pod husk extract gel on day 7 and 14. *Significant p values (p ≤ 0.05).
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