Selective blockade of voltage-gated potassium channels reduces inflammatory bone resorption in experimental periodontal disease.
The effects of the potassium channel (Kv1.3) blocker kaliotoxin on T-cell-mediated periodontal bone resorption were examined in rats. Systemic administration of kaliotoxin abrogated the bone resorption in conjunction with decreased RANKL mRNA expression by T-cells in gingival tissue. This study suggests a plausible therapeutic approach for inflammatory bone resorption by targeting Kv1.3.


BACKGROUND
Kv1.3 is a critical potassium channel to counterbalance calcium influx at T-cell receptor activation. It is not known if Kv1.3 also regulates RANKL expression by antigen-activated T-cells, and consequently affects in vivo bone resorption mediated by activated T-cells.


METHODS
Actinobacillus actinomycetemcomitans 29-kDa outer membrane protein-specific Th1-clone cells were used to evaluate the expression of Kv1.3 (using reverse transcriptase-polymerase chain reaction [RT-PCR] and Western blot analyses) and the effects of the potassium channel blocker kaliotoxin (0-100 nM) on T-cell activation parameters ([3H]thymidine incorporation assays and ELISA) and expression of RANKL and osteoprotegerin (OPG; flow cytometry, Western blot, and RT-PCR analyses). A rat periodontal disease model based on the adoptive transfer of activated 29-kDa outer membrane protein-specific Th1 clone cells was used to analyze the effects of kaliotoxin in T-cell-mediated alveolar bone resorption and RANKL and OPG mRNA expression by gingival T-cells. Stimulated 29-kDa outer membrane protein-specific Th1 clone cells were transferred intravenously on day 0 to all animals used in the study (n = 7 animals per group). Ten micrograms of kaliotoxin were injected subcutaneously twice per day on days 0, 1, 2, and 3, after adoptive transfer of the T-cells. The control group of rats was injected with saline as placebo on the same days as injections for the kaliotoxin-treated group. The MOCP-5 osteoclast precursor cell line was used in co-culture studies with fixed 29-kDa outer membrane protein-specific Th1-clone cells to measure T-cell-derived RANKL-mediated effects on osteoclastogenesis and resorption pit formation assays in vitro. Statistical significance was evaluated by Student's t-test.


RESULTS
Kaliotoxin decreased T-cell activation parameters of 29-kDa outer membrane protein-specific Th1 clone cells in vitro and in vivo. Most importantly, kaliotoxin administration resulted in an 84% decrease of the bone resorption induced in the saline-treated control group. T-cells recovered from the gingival tissue of kaliotoxin-treated rats displayed lower ratios of RANKL and OPG mRNA expression than those recovered from the control group. The ratio of RANKL and osteoprotegerin protein expression and induction of RANKL-dependent osteoclastogenesis by the activated T-cells were also markedly decreased after kaliotoxin treatments in vitro.


CONCLUSIONS
The use of kaliotoxin or other means to block Kv1.3 may constitute a potential intervention therapy to prevent alveolar bone loss in periodontal disease.
Valverde
Paloma
P
Department of Oral and Developmental Biology, Harvard Medical School and Harvard School of Dental Medicine, Boston, Massachusetts, USA. paloma.valverde@tufts.edu
Kawai
Toshihisa
T
Taubman
Martin A
MA
D000135:Acid Phosphatase / Q000032:analysis; D019264:Adoptive Transfer; D016976:Aggregatibacter actinomycetemcomitans / Q000737:chemistry; D016301:Alveolar Bone Loss / Q000188:drug therapy* / Q000276:immunology / Q000503:physiopathology; D000818:Animals; D017951:Antigen Presentation / Q000187:drug effects / Q000276:immunology; D001425:Bacterial Outer Membrane Proteins / Q000276:immunology / Q000494:pharmacology; D015153:Blotting, Western; D017252:CD3 Complex / Q000276:immunology; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects / Q000276:immunology; D018920:Coculture Techniques; D015536:Down-Regulation / Q000276:immunology / Q000502:physiology; D005260:Female; D015870:Gene Expression; D006023:Glycoproteins / Q000235:genetics / Q000378:metabolism; D007074:Immunoglobulin G / Q000097:blood; D007371:Interferon-gamma / Q000378:metabolism; D007527:Isoenzymes / Q000032:analysis; D051664:Kv1.3 Potassium Channel; D008070:Lipopolysaccharides / Q000494:pharmacology; D008213:Lymphocyte Activation / Q000187:drug effects; D008437:Maxilla / Q000473:pathology; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism; D010010:Osteoclasts / Q000276:immunology / Q000502:physiology; D053244:Osteoprotegerin; D010518:Periodontitis / Q000188:drug therapy* / Q000276:immunology / Q000503:physiopathology; D015221:Potassium Channels / Q000187:drug effects / Q000235:genetics / Q000502:physiology*; D024642:Potassium Channels, Voltage-Gated / Q000037:antagonists & inhibitors; D053245:RANK Ligand; D051381:Rats; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics / Q000378:metabolism; D018124:Receptors, Tumor Necrosis Factor; D012604:Scorpion Venoms / Q000494:pharmacology*; D013154:Spleen / Q000166:cytology; D013601:T-Lymphocytes / Q000276:immunology / Q000637:transplantation; D000071681:Tartrate-Resistant Acid Phosphatase
D001425:Bacterial Outer Membrane Proteins; D017252:CD3 Complex; D002352:Carrier Proteins; D006023:Glycoproteins; D007074:Immunoglobulin G; D007527:Isoenzymes; C496975:Kcna3 protein, rat; D051664:Kv1.3 Potassium Channel; D008070:Lipopolysaccharides; D008562:Membrane Glycoproteins; C117494:Omp29 protein, Actinobacillus actinomycetemcomitans; D053244:Osteoprotegerin; D015221:Potassium Channels; D024642:Potassium Channels, Voltage-Gated; D053245:RANK Ligand; D018160:Receptors, Cytoplasmic and Nuclear; D018124:Receptors, Tumor Necrosis Factor; D012604:Scorpion Venoms; C506032:Tnfrsf11b protein, rat; C072276:kaliotoxin; D007371:Interferon-gamma; D000135:Acid Phosphatase; D000071681:Tartrate-Resistant Acid Phosphatase