Experimental gingivitis in young dogs

abstract — The aim of the present study was to analyze and express in quantitative terms some of the structural alterations which develop in an initially normal gingiva during a phase of continuous plaque accumulation. Four beagle dogs were used. The animals had from birth been twice daily subjected to meticulous mechanical tooth cleaning. When the dogs were 10 months of age their gingiva were in excellent health as evaluated by Gingival Index and Gingival Exudate measurements. Gingival tissues were harvested from the premolar and molar regions in the right jaws. The tooth cleanings were then terminated and plaque allowed to accumulate. Clinical examinations were performed and gingival biopsies sampled after 4, 7, 14, 21 and 28 d. The composition of the gingival biopsies was analyzed in a sampling microscope. After 4 d of plaque accumulation significant amounts of gingival exudate could be sampled. The exudation then gradually increased during the following weeks. Biopsies representing day zero did not contain any inflammatory cell infiltrates. However, after 4 d of the experiment leukocytes were found in the collagen-poor connective tissue immediately beneath the junctional epithelium. The size of the infiltrated connective tissue (ICT) gradually increased during the experiment. The volumetric density of collagen in the noninfiltrated connective tissue (NCT) was always much higher than in ICT. In ICT, however, this density parameter remained rather constant throughout the study. On days 4 and 7 neutrophilic granulocytes constituted 60–70% of the leukocyte population. On day 28, however, the infiltrate comprised mainly mononuclear leukocytes, especially plasma cells, neutrophils at that time occupying only a small fraction of the infiltrate.     

Experimental gingivitis in immunized dogs

The present experiment was performed to study the clinical and histopathological features of developing gingivitis in eight dogs, of which four were repeatedly immunized with plaque antigens prior to the start of a defined plaque accumulating period. The dogs were, after weaning, fed a soft diet but also twice daily subjected to meticulous tooth cleaning. When they were 10 months old and their gingiva were in excellent health, all tooth cleaning procedures were terminated and dental plaque was allowed to accumulate for 4 weeks. Clinical examinations and gingival biopsy sampling were performed during this period. The biopsies were analyzed morphometrically.
In the injected dogs the levels of specific serum antibodies, as determined by an immunoradiometric assay (IRMA), were raised 3–4 fold. No antibody increase was observed in the controls. After the termination of the tooth-cleaning period, the immunized dogs seemed to accumulate dental plaque at the same rate as non-immunized controls. The gingiva of the immunized animals reacted to initial plaque formation with symptoms such as redness, swelling and bleeding in a manner very similar to the controls. The degree of gingival exudation and the size of the gingival connective tissue lesion was, however, less pronounced in the immunized dogs compared to the non-immunized controls.
The results indicate that raised immunity to plaque antigens may have a protective function in the dento-gingival tissues during the development of plaque induced gingivitis.     

Necrotizing ulcerative gingivitis in beagle dogs

The induction and transmission of Necrotizing Ulcerative Gingivitis (NUG) lesions in beagle dogs have been reported previously. In the present experiments the contagious nature of these NUG lesions was further investigated by studying the unilateral induction and the intraoral transmission of NUG lesions. Six beagle dogs were treated with Kenacort A40®. The control quadrants of their dentition were cleaned while the experimental quadrants were not. The experimental quadrants were inoculated with pooled dental debris from the same dogs. NUG lesions were induced in the experimental quadrants, but NUG lesions were observed also at some sites of the control quadrants. In the second part of the experiment, reinoculation of non-affected experimental sites with debris from lesions of the same dog or cage companion increased the number of NUG lesions in both experimental and control quadrants.
The study of experimental intraoral transmission of NUG lesions was restricted to the non-diseased sites in experimental and control quadrants. Topical application of debris of NUG lesions on the test sites did not spread the disease to these sites. Microscopic and cultural evaluation of subgingival microflora revealed no differences in the presence of Bacteroides, Fusobacteria, Spirocheata , and Spirillum in relation to the presence or absence of NUG. Three different types of black pigmental Bacteroides species were observed. It is concluded, that the morphology of the interdental sites is of great importance for the clinical occurence of NUG lesions and that the increase in number and severity of lesions after local reinoculation indicates that in the phase of primary infection, iatrogenic NUG lesions might be possible in a susceptible host.     

Necrotizing ulcerative gingivitis in beagle dogs

A new modification of the Warthin spirochete staining method was developed to study at the light microscopic level the distribution of spirochetes in gingival tissues of beagle dogs with induced necrotizing ulcerative gingivitis. Spirochetes were present in cellular exudate at ulcerated sites, in edematous epithelium in the vicinity of ulcerations, and in intact sulcular epithelium. They did not appear to penetrate connective tissues via the ulcerations. These observations suggest that, in the etiology of NUG, spirochetes play a role in the initiation of the pathologic process by penetration of the intercellular spaces in the epithelium.     

Necrotizing ulcerative gingivitis in beagle dogs

Clinical studies in beagle dogs have indicated that NUG occurs at sites of predilection. The purpose of the present investigation was to compare the histologic characteristics of interdental sites exhibiting clinical NUG with those without clinical NUG in relation to the presence or absence of interproximal contacts.
In six beagle dogs, aged three years, NUG was experimentally induced. At the end of a longitudinal clinical study the dogs were sacrified and prepared for histological evaluation. The occurrence of ulcerations in the col epithelium and the adjacent epithelium is related to the clinical NUG scores as well as to the presence of interproximal contact. The same holds true for the presence of an inflammatory infiltrate in the subjacent connective tissue. Ulcerations of the col epithelium as well as the infiltrate subjacent to this area appears to be related to the clinical NUG scores, but the ulcerations in the sulcular and junctional epithelium are not, nor is the infiltrate subjacent to these epithelial areas.
The results suggest two possibilities. The first is that the clinical NUG score is insufficient to trace the sulcular expression of the disease, and the second possibility is the existence of two different periodontal diseases in these beagle dogs.     

Vascular permeability during initial gingivitis in dogs

Variations in the permeability of dento-gingival blood vessels during initial gingivitis have been studied in four beagle dogs. On two occasions during a four-day period of developing gingivitis, increased permeability of the dento-gingival blood vessels was provoked by inserting filter paper strips into the dento-gingival junction. Vessels in a state of abnorma permeability were labelled on one occasion during the four-day period by intravascularly injected colloidal carbon and by fluorescent microspheres on the other occasion. The vascular labelling scores for each of the two tracers were determined in gingival biopsies. An experimental outline was used which allowed comparison of the permeability characteristics of the vessels on day 0 with those on day 2 and on day 2 with those on day 4 of the initial gingivitis.
At each time interval (day 0, 2 and 4) the vascular labelling of the dento-gingival blood vessels was found evenly distributed in the coronal-apical direction of the vascular plexus. The labels were most frequently found in vessels 10–50 microns in width. In three of the dogs the vascular label scores were found to be significantly higher on day 2 compared with day 0 and on day 4 compared with day 2.
The results indicate the development of a vasculitis in the dento-gingival blood vessels during initial gingivitis involving most vessels of venular type in the vascular plexus.     

Sulfadiazines reduce gingivitis and plaque formation in beagle dogs

The effect of zinc sulfadiazine (ZnSD) and silver sulfadiazine (AgSD) on reducing plaque formation and gingivitis was studied in 12 beagle dogs over a 14-week period. 12 beagle dogs were scaled, root planed and pumiced to bring them to a similar level of gingival health, prior to placing them on a diet of Purina Dog Chow softened with canned gravy and molasses to promote the build-up of plaque and the initiation of gingivitis. At the end of 8 weeks, the dogs were determined to have substantial bacterial plaque accumulation and apparent gingivitis. Thereafter, 4 dogs were treated 2 x daily with topical applications of 3% zinc sulfadiazine; 4 dogs were treated with 2% silver sulfadiazine while 4 dogs were treated with placebo gel serving as control over a 14-week treatment period. By week 2, the zinc and silver sulfadiazine dogs showed a significant decrease in gingival index which was maintained throughout the study. Additionally, by week 2, the % of sites with bleeding was also seen to decrease significantly in the experimental groups. The plaque index remained consistent in all 3 groups until week 6 when the 2 experimental groups indicated significant decrease in plaque accumulation as compared to controls. Probing depths were also seen to decrease significantly in the experimental groups after 10 weeks of therapy. The mean stain index was similar in all 3 groups of dogs throughout the study. Data indicate that both zinc and silver sulfadiazine inhibit plaque formation and reduce existing gingivitis in beagle dogs.     

Effect of experimental neutropenia on initial gingivitis in dogs

Abstract – The role of neutrophilic granulocytes in the loss of gingival collagen has been studied by inducing experimental neutropenia during initial gingivitis in beagle dogs. Neutropenia was induced for 4 d in three animals with normal gingiva by repeated injections of rabbit anti-neutrophil serum. During neutropenia microbial plaque was allowed to form on the teeth. Samples of junctional (crevicular) leukocytes and gingival fluid were taken on days 0 and 4. Block biopsies of buccal gingiva were obtained on day 4. Stained semi- and ultrathin sections were used for histometric and serologic tissue analysis. Gingival fluid flow increased from day 0 to day 4 in all dogs while junctional leukocytes increased in one dog only. Subgingival plaque had formed in most biopsies, and in the junctional epithelium very few neutrophilic granulocytes were present. In the coronal connective tissue subjacent to the junctional epithelium lymphoid cells, structurally abnormal neutrophilic granulocytes and monocytes/macrophages were diffusely scattered. The gingival collagen appeared mainly displaced by the inflammatory cells rather than dissolved. The data suggest that neutrophilic granulocytes may contribute to the loss of gingival collagen during initial gingivitis in dogs. The neutrophils also seem to be of importance for the limitation of subgingival plaque growth along the tooth surface.     

Influence of experimental neutropenia in dogs with chronic gingivitis

The influence of a four day period of experimental neutropenia on the clinical state and the structural constituents of chronically inflamed gingiva has been studied in four beagle dogs. Neutropenia was induced by heterologous anti-neutrophil serum. The effects on the gingiva were evaluated by Gingival Index (GI) and Gingival Exudate measurements and by morphometric analysis of various tissue components in sections from biopsies of buccal gingiva sampled on days 2, 3, and 4. Control biopsies were obtained before induction of neutropenia. The GI did not markedly change during the observation period. The amount of Gingival Exudate, however, significantly decreased following the induction of neutropenia and remained low throughout the experiment. Neutrophilic granulocytes disappeared in the gingival tissues, while, apart from a relative increase in plasma cells, no other tissue components seemed to change. No bacterial invasion of the junctional epithelium or of the gingival connective tissue could be observed. Thus during a four day period the bacterial defense mechanisms at the dentogingival junction seem to be able to prevent penetration of microorganisms into the gingival tissues despite the absence of neutrophilic granulocytes.     

The role of keratinized gingiva in plaque-associated gingivitis in dogs

Abstract The present experiment was designed to examine if a dento-gingival unit with a wide zone of keratinized gingiva (KG) provides a more efficient seal against plaque infection than a unit with a narrow zone of KG or lack of such gingiva. Five beagle dogs were used. In a preparatory experiment three categories of gingival units were established in each dog, namely (1) regenerated gingiva with a wide zone of KG (healed following a “flap” procedure), (2) regenerated gingiva with an “insufficiently” wide zone of KG (healed following a “gingivectomy” procedure), and (3) non-operated control gingiva with a normal zone of KG. Clinical and histological examinations carried out at the end of the preparatory period revealed that all gingival units, non-operated as well as regenerated units, met the criteria of health. All tooth-cleaning measures were abolished. After 40 days of plaque accumulation the clinical examination (Plaque Index, Gingival Index and Gingival Exudate) was repeated and biopsies were sampled. Following preparation the biopsy material was subjected to histometric and morphometric analysis. The results showed that the free gingiva of units with wide and narrow zones of KG responded to microbial colonization by an inflammatory reaction, the location and extension of which did not vary with the width of the keratinized gingiva. It was concluded that, in the presence of plaque, a dento-gingival unit with a narrow zone of, or with lack of, gingiva has an equal capacity for inflammatory response against plaque infection as a unit with a wide zone of KG.     

Development of plaque and gingivitis following antibiotic therapy in dogs

Abstract Changes in the subgingival microbiota were studied during a 4-week period of plaque accumulation in beagles which initially had clean teeth and healthy gingiva. The effect of Clindamycin, Metronidazole and Vancomycin on the microbiota and associated gingivitis was also determined. Fifteen 1-year-old beagle dogs were used. During two subsequent periods of plaque accumulation, one without (control) and one with (test) antibiotic therapy, the animals were allowed to accumulate plaque. Plaque and gingivitis development was assessed by clinical measurements. Plaque samples were harvested from the gingival sulcus region and the percentage distribution of different bacteria was determined following aerobic and anaerobic incubation in selected media. Following microbial sampling, biopsies of the gingiva and surrounding soft and hard tissues were removed and the size of the inflammatory cell infiltrate assessed. The results demonstrated that beagle dogs with a normal gingiva harbored a microbiota within the gingival sulcus region dominated by gram-positive and gram-negative rods. After 4 weeks of undisturbed plaque accumulation, the number of sulcus bacteria had increased 100-fold. This increase was the result of a proliferation of gram-negative anaerobic rods. Concomitant with this change of the microflora, an inflammatory reaction developed in the gingiva. When the dogs during a similar 4-week period were given Metronidazole, no increase of the subgingival microbiota occurred. The inflammatory response of the gingiva was minute. Treatment with Clindamycin and Vancomycin did not entirely prevent the proliferation of the subgingival bacteria. The number of microorganisms harbored after 4 weeks of Clindamycin and Vancomycin treatment was, however, only 50% of that of the control period. Even if the number of colony forming units was similar at the end of Vancomycin and Clindamycin treatment the quality of the two subgingival microbiotas was different. Vancomycin treatment resulted in a plaque dominated by gram-negative bacteria while Clindamycin allowed gram-positive bacteria to increase in number. Gingival inflammation resulting in the Vancomycin group was more pronounced than the gingivitis response in the Clindamycin group.     

Inhibition of experimental gingivitis in beagle dogs with topical mercaptoalkylguanidines

BACKGROUND: Nitric oxide is a free radical produced in host tissues by constitutive and inducible forms of the enzyme nitric oxide synthase. Nitric oxide plays physiological roles, but it is also involved in the pathophysiology of several inflammatory conditions, including arthritis, ulcerative colitis, and circulatory shock. Local increases in inducible nitric oxide synthase (iNOS) and reactive nitrogen products have also been demonstrated in humans and animals with periodontal disease. This masked, randomized, placebo-controlled preclinical investigation examined the effect of two mercaptoalkylguanidines, mercaptoethylguanidine (MEG) and guanidinoethyldisulfide (GED), which are iNOS inhibitors and reactive nitrogen scavenging compounds, on the development of experimental gingivitis in beagle dogs. METHODS: Fifteen female, 1-year-old beagles first completed a 2-week dose-escalation experiment during which a maximum tolerated dose was determined for MEG and GED gels. Thereafter, all animals were brought to optimal gingival health by mechanical scaling, followed by rigorous daily toothbrushing over a 4-week washout period. Experimental gingivitis was then induced, with cessation of plaque control and institution of a soft diet over 8 weeks. Beagles randomly received 0.3% MEG, 0.3% GED, or placebo (vehicle) gels, topically applied twice daily to premolar teeth. Gingival inflammation, bleeding tendency, and supragingival plaque were clinically measured at baseline and at 2, 3, 4, 6, and 8 weeks. Comparisons among groups and between group pairs (active versus placebo) were made using Kruskal-Wallis tests. RESULTS: From baseline to day 7, all groups expressed similar indices. Thereafter, significant and time-dependent increases in the plaque index (PI), gingival index (GI), and percentage of bleeding on probing (%BOP) were observed in placebo-treated beagles. Mean GI scores for beagles treated with GED or MEG gels remained at or below baseline levels for the entire treatment period. At weeks 2, 3, 4, and 8, GI scores were significantly lower for MEG and GED groups compared to the placebo group (P<0.05). In addition, MEG and GED gels significantly reduced gingival bleeding responses by 8 weeks (P<0.05). Although placebo-treated beagles demonstrated %BOP scores of 43% at week 8, GED- and MEG-treated beagles exhibited %BOP scores of 21% and 26%, respectively. Since no statistical difference among PI scores was noted for any of the time points, neither mercaptoalkylguanidine appeared to affect supragingival plaque levels. CONCLUSION: The data from this preclinical study indicate that mercaptoalkylguanidines, topically administered, may significantly reduce experimental gingivitis in the beagle dog.     

Initial gingivitis induced by topical application of plaque extract

The present study was undertaken to examine if soluble substances in human dental plaque were able to induce structural alterations in the marginal gingiva characteristic of initial gingival inflammation.
Four-day old plaque was sampled from dental students; the plaque material was pooled, suspended in saline, homogenized, centrifuged and filtered. The supernatant, the plaque-extract, was then, during an eight hour period, topically applied to the gingival margin of the upper jaws of five Beagle dogs. In control regions saline was applied in an identical manner. One and a half hour before sacrifice the animals were given i.v. injections of colloidal carbon.
Biopsies of test and control regions were taken according to a technique described by Schroeder et al (1973). The biopsy specimens were embedded in Epon. Semithin sections (2-3 μm) were examined in the microscope regarding 1. amount of vascular units subjacent to the junctional epithelium containing carbon particles
2. number of polymorphonuclear and mononuclear leucocytes within the junctional epithelium.
The examinations revealed that the gingival units exposed to plaque extract in comparison to controls harboured larger numbers of carbon labelled vascular units and larger numbers of leukocytes within the junctional epithelium. The data also showed that the main reaction occurred in the marginal portion of the gingival tissue. The observations of this study indicate that soluble substances from human dental plaque when applied topically in the gingival sulcus may give rise to structural alterations similar to those developing in the gingival tissues in response to plaque accumulation.     

Effect of anti-thymocyte serum on development of gingivitis in dogs

The effect of repeated injections of heterologous anti-thymocyte globulin (ATS) on the development of gingivitis has been studied in beagle dogs. Control dogs were injected with control rabbit immunoglobulin. The immunosuppressive effect of the antiserum was evaluated from the inhibition of a delayed hypersensitivity reaction in the skin towards l-dinitro-2.4-chlorobenzene (DNCB). The gingival condition was determined by measurements of crevicular leukocytes, gingival fluid, acid phosphatase activity in crevicular samples and by counts of inflammatory cells in sections of gingival biopsies. Antiserum administration, but not control rabbit IgG, abolished the hypersensitivity reaction in the skin towards DNCB. The parameters of gingival inflammation increased during the 28day experimental period, both in the experimental and in the control dogs. The results indicate that cellular immunity does not play a major role for the development of gingival inflammation in beagle dogs.     

云南白药对比格犬牙龈炎的治疗作用

目的:评价云南白药对比格犬牙龈炎的治疗作用。方法:12只符合条件的比格犬随机分为云南白药组和空白对照组。2组动物分别用浸有1.23g/L云南白药溶液或生理盐水的棉球敷于牙龈治疗,1min/次,2次/d,持续14d。在给药前和末次给药后24h分别盲法对动物的菌斑(P),牙龈炎症状(G)和炎性浸润(INF)评分。结果:空白对照组动物的P、G、INF没有显著性差异(P>0.1),云南白药组动物的各评分指标均有显著性改善(P<0.05)。结论:云南白药可抑制比格犬牙龈炎菌斑形成,减轻牙龈炎症状和炎细胞的聚集。实验结果提示云南白药可能对临床牙龈炎有一定治疗作用。     

Morphological changes in gingival blood vessels during developing gingivitis in dogs

Blood vessels situated at the gingival margin were studied in dogs during a 16-day period of developing gingivitis. Morphological changes in the vessels were assessed from photographs which had been taken periodically and were identical. Two characteristic alterations were found: increased width and changes in the course of the vessels. Increases in width were observed during the first week of the experiment. Changes in the course of vessels were found during the second week.     

Inhibition of experimental gingivitis in beagle dogs with topical salivary histatins

Abstract Histatins, histidine-rich proteins found within parotid and submandibular secretions, are a novel class of endogenous peptides with antimicrobial properties. This masked, randomized, placebo-controlled preclinical investigation examined the effect of 3 topical histatins on the development of plaque and gingivitis in beagle dogs. 16, female, 1-year-old beagles were brought to optimal gingival health by mechanical scaling and polishing followed by rigorous daily tooth brushing. At the conclusion of this pretreatment period, dogs were randomly divided into 4 groups for the application of test formulations, and were placed on a plaque-promoting diet, Test agents included 3 synthetic salivary histatins (histatin 5, P-113 and P-113D) which were incorporated in hydroxypropyl methylcellulose gel at a concentration of 0.125%, and a placebo, or negative control, which was the gel vehicle alone. Throughout the 10-week treatment period, test formulations (2.0 ml) were applied 2xdaily to all premolar teeth using a Monojet syringe. Plaque formation and gingival inflammation were assessed using the plaque (PI) and gingival (GI) indices on days 0, 7, 14, 21, 28, 42, 56 and 70. Furthermore, bleeding to probing was recorded as a percent of sites (%BOP) and according to the modified sulcus bleeding index (mSBI). Comparisons among groups and between group pairs (active versus placebo) were made with Kruskal-Wallis tests with the average of data over the interval, days 14–42, being the primary focus of the analysis. From baseline to day 7, all groups expressed similar indices. Thereafter, overall significant differences among the groups were noted at day 42 for PI, at days 21, 28, 42 and 70 for GI, and at days 14 and 28 for %BOP (p<0.05). In particular, beagles treated with P-113 demonstrated significantly lower PI scores at day 42 (p<0.05), significantly lower GI scores from days 21 through 42 (<0.05), and significantly lower %BOP scores at days 14 and 28 (p<0.05) compared to beagles treated with placebo. Beagles treated with P-113D exhibited significantly lower GI at day 42 compared to the placebo (p<0.05). For the primary analysis conducted over the midtreatment interval (days 14–42), significant differences were detected for all parameters except mSBI (p<0.05). Accordingly, significantly lower PI scores were found for P-113, lower GI scores for P-113 and P-113D, and lower %BOP for P-113 and P-113D compared to placebo (p<0.05). These data indicate that in the beagle model, salivary histatins, P-113 and P-113D, topically applied, can significantly reduce clinical signs of plaque formation and gingival inflammation.     

Lymphoproliferative response of submandibular nodes to mitogens in experimental gingivitis in dogs

The purpose of the present study was to evaluate the lymphoproliferative response of submandibular nodes cells to stimulation by phytohemagglutinin (PHA), concanavalin A (Con A), and lipopolysaccharide (LPS) following experimental gingivitis in dogs.
In 4 dogs of the experimental group, gingival inflammation was induced by cervical ligatures on the left jaw, and the right jaw was kept clean by toothbrushing. In 2 dogs of the control group, toothbrushing was performed throughout. After the weight and lymphocyte number in the submandibular nodes were calculated, the lymphocyte response was determined by the uptake of ³H-thymidine.
The weight and lymphocyte number of the ipsilateral nodes increased 2.2 and 3.7 times, respectively, as compared with the contralateral nodes. There was a higher background proliferation of unstimulated lymphocytes from ipsilateral nodes. The lymphocyte response to lower PHA concentration from ligated side lymphnodes significantly increased compared to that from contralateral nodes. It is suggested that the plaque accumulation and the intensive gingival inflammation considerably influence lymphocytes in the submandibular nodes.     

The appearance of bone resorbing activity in ligature induced gingivitis in dogs

The in vitro bone resorbing activity of extracts from healthy and inflamed dog gingiva were compared and the appearance of the resorbing factor between 1 and 6 weeks of gingivitis was measured. Seven adult dogs were used. The gingiva were initially rendered healthy by scaling and polishing. Plaque retentive ligatures were placed around the necks of teeth so that at the end of the experiment samples of healthy gingiva and gingiva with 1 and 6 week old ligature induced gingivitis could be harvested from each dog. The bone resorbing activity of extracts from these gingival samples was studied by measuring the release of ⁴⁵Ca from prelabeled fetal rat long bones in organ culture. Six out of 7 extracts from healthy quadrants had no bone resorbing potential whereas 13 out of 14 extracts from ligatured quadrants stimulated significant amounts of bone resorption. The resorbing activity appeared during the first week after ligature placement and no significant increase in resorbing potential was noted between extracts obtained 1 and 6 weeks after ligature placement. It was concluded that the appearance of the bone resorbing factor(s) is related to the accumulation of plaque and/or the resultant inflammation and that it may be responsible for the alveolar bone loss seen in ligature induced periodontal disease in dogs.