A single session of antimicrobial photodynamic therapy does not influence the alveolar repair process in rats.

The aim of this study was to use microscopic and molecular techniques to evaluate the effects of a single session of antimicrobial photodynamic therapy (aPDT) on the alveolar repair process after tooth extraction in rats. The study sample included 84 rats divided into four groups, as follows: a) Control - untreated socket; b) Laser - socket treated using photobiomodulation; c) TBO - socket treated with topic application of the photosensitizer agent, toluidine blue O (TBO); and d) aPDT - socket treated with TBO and laser irradiation. An additional rat was used for thermal mapping during socket irradiation. The animals were euthanatized at 6, 15, and 28 days after unilateral extraction of the upper incisor. Quantitative and qualitative analyses of the connective and bone tissues, blood clot, blood vessel, and inflammatory infiltrate were performed, and real-time polymerase chain reaction was used to study the expression of genes (collagen type I, osteocalcin, alkaline phosphatase [ALP], runt-related transcription factor 2 [RUNX2], and vascular endothelial growth factor [VEGF]) involved in the bone healing process. No statistically significant differences in microscopic and molecular outcomes were observed between the groups (p > 0.05). A positive correlation was seen to exist between blood clot and VEGF (p = 0.000), and a negative correlation was observed between bone tissue and ALP (p = 0.028) and blood vessel and VEGF (p = 0.018). A single session of aPDT in the dental extraction site did not influence the alveolar repair process in rats.

A single session of antimicrobial photodynamic therapy does not influence the alveolar repair process in rats

Abstract: The aim of this study was to use microscopic and molecular techniques to evaluate the effects of a single session of antimicrobial photodynamic therapy (aPDT) on the alveolar repair process after tooth extraction in rats. The study sample included 84 rats divided into four groups, as follows: a) Control - untreated socket; b) Laser - socket treated using photobiomodulation; c) TBO - socket treated with topic application of the photosensitizer agent, toluidine blue O (TBO); and d) aPDT - socket treated with TBO and laser irradiation. An additional rat was used for thermal mapping during socket irradiation. The animals were euthanatized at 6, 15, and 28 days after unilateral extraction of the upper incisor. Quantitative and qualitative analyses of the connective and bone tissues, blood clot, blood vessel, and inflammatory infiltrate were performed, and real-time polymerase chain reaction was used to study the expression of genes (collagen type I, osteocalcin, alkaline phosphatase [ALP], runt-related transcription factor 2 [RUNX2], and vascular endothelial growth factor [VEGF]) involved in the bone healing process. No statistically significant differences in microscopic and molecular outcomes were observed between the groups (p > 0.05). A positive correlation was seen to exist between blood clot and VEGF (p = 0.000), and a negative correlation was observed between bone tissue and ALP (p = 0.028) and blood vessel and VEGF (p = 0.018). A single session of aPDT in the dental extraction site did not influence the alveolar repair process in rats.

Acinar autolysis and mucous extravasation in human sublingual glands: a microscopic postmortem study

Although some morphological investigations on aged human sublingual glands (HSG) found eventual phenomena identified as autolysis and mucous extravasation, the exact meaning of these findings has not been elucidated.Objective The aim of this work is to investigate whether acinar autolysis and mucous extravasation are related to the aging process in human sublingual glands. We also speculate if autolytic changes may assist forensic pathologists in determining time of death.Material and Methods 186 cadavers’ glands were allocated to age groups: I (0–30 years); II (31–60), and III (61–90). Time and mode of death were also recorded. Acinar autolysis and mucous extravasation were classified as present or absent. Ultrastructural analysis was performed using transmission electron microscopy (TEM). Data were compared using Mann-Whitney U, Spearman’s correlation coefficient, Kruskal-Wallis, and Dunn tests (p<0.05).Results There was correlation between age and acinar autolysis (r=0.38; p=0.0001). However, there was no correlation between autolysis and time of death. No differences were observed between genders. TEM showed mucous and serous cells presenting nuclear and membrane alterations and mucous cells were more susceptible to autolysis.Conclusion Acinar autolysis occurred in all age groups and increased with age while mucous extravasation was rarely found. Both findings are independent. Autolysis degrees in HSG could not be used to determine time of death.

Assessment of new bone formation around titanium surface treated implants in diabetic rats / Neoformação óssea ao redor de implantes de titânio com superfície tratada inseridos em ratos diabéticos

Objective: This study aimed to compare the bone formation around titanium implants with machined and acid-etched surfaces, inserted in induceddiabetic rats and in non-diabetic rats, in an attempt to investigate whether there are differences in bone formation between this metaboliccondition and the use of different implant surfaces. Methods: Custom fabricated commercially pure solid cylinder titanium implants, machined and acid-etched surface were inserted in the femora of streptozotocin-induced diabetic rats (n=10) and non-diabetic rats (n=10). Morphometrical bone-implant contact percentage and bone area within the limits of the implant threads (BD) were performed at 21 days of healing. Results: Peri-implant tissue in machined implant showed intense new bone formation within all threads of the implants of the non-diabetic group (BIC= 82.8 ± 9.23 e BD = 38.7 ± 4.27) while diabetic group (BIC = 35.3 ± 9.4 and BD = 20.0 ± 3.8) exhibited small and immature bone formationwithin threads of the implants with thickness fibrous connective tissue interposition between bone-implant interface. In the acid-etchedsurface implants in both, diabetic and non-diabetic groups, the peri-implant tissue showed intense new bone formation within all threads ofthe implants with BIC = 74.4 ± 14.7 e BD = 35.4 ± 3.48 in non-diabetic group and BIC = 63.1 ± 12.9 e BD = 29.6 ± 4.9 in diabetic group.Conclusion: In machined surface implants the diabetes interfere negatively in osseointegration while acid-etched surface promoted major BIC and BD index, indicating its selective use in diabetic patients.

Objetivo: Comparar a formação óssea ao redor de implantes de superfície lisa e tratada, instalados em ratos diabético-induzidos e não-diabéticos,investigando se há diferenças na formação óssea entre os dois quadros metabólicos, melhora no padrão de osteogênese entre as diferentessuperfícies e, sua relação com o diabetes. Métodos: Foram instalados implantes de titânio de superfícies lisa e tratada, no fêmur de ratos diabético-induzidos com estreptozotocina (n=10) e não diabéticos (n=10).A Análise morfométrica da porcentagem de contato osso-implante (COI) foi realizada 21 dias após a cirurgia. Resultados: A neoformação óssea foi intensa ao redor dos implantes de superfície lisa nos ratos não-diabéticos (COI = 82.8 ± 9.23), enquanto que o grupo diabético exibiu pequena e imatura formação óssea (COI=35.3 ± 9.4), com interposição de tecido conjuntivo na interface osso-implante. Ao redor dos implantes com superfície tratada, ocorreu intensa neoformação óssea, tanto nos animais diabéticos (COI = 63.1 ± 12.9) como nos não-diabéticos, (COI = 74.4 ± 14.7). Conclusão: Nos implantes de superfície lisa, o diabetes interfere negativamente na osseointegração, enquanto que as superfícies tratadas com ácido promoveram maior contato osso-implante, indicando seu uso seletivo em pacientes diabéticos.

Biocompatibility and setting time of CPM-MTA and white Portland cement clinker with or without calcium sulfate.

OBJECTIVE: To evaluate the biocompatibility and the setting time of Portland cement clinker with or without 2% or 5% calcium sulfate and MTA-CPM. MATERIAL AND METHODS: Twenty-four mice (Rattus norvegicus) received subcutaneously polyethylene tubes filled with Portland cement clinker with or without 2% or 5% calcium sulfate and MTA. After 15, 30 and 60 days of implantation, the animals were killed and specimens were prepared for microscopic analysis. For evaluation of the setting time, each material was analyzed using Gilmore needles weighing 113.5 g and 456.5 g, according to the ASTM specification Number C266-08 guideline. Data were analyzed by ANOVA and Tukey's test for setting time and Kruskal-Wallis and Dunn test for biocompatibility at 5% significance level. RESULTS: Histologic observation showed no statistically significant difference of biocompatibility (p>0.05) among the materials in the subcutaneous tissues. For the setting time, clinker without calcium sulfate showed the shortest initial and final setting times (6.18 s/21.48 s), followed by clinker with 2% calcium sulfate (9.22 s/25.33 s), clinker with 5% calcium sulfate (10.06 s/42.46 s) and MTA (15.01 s/42.46 s). CONCLUSIONS: All the tested materials showed biocompatibility and the calcium sulfate absence shortened the initial and final setting times of the white Portland cement clinker.

Biocompatibility and setting time of CPM-MTA and white Portland cement clinker with or without calcium sulfate

Objective: To evaluate the biocompatibility and the setting time of Portland cement clinker with or without 2% or 5% calcium sulfate and MTA-CPM. Material and Methods: Twenty-four mice (Rattus norvegicus) received subcutaneously polyethylene tubes filled with Portland cement clinker with or without 2% or 5% calcium sulfate and MTA. After 15, 30 and 60 days of implantation, the animals were killed and specimens were prepared for microscopic analysis. For evaluation of the setting time, each material was analyzed using Gilmore needles weighing 113.5 g and 456.5 g, according to the ASTM specification Number C266-08 guideline. Data were analyzed by ANOVA and Tukey's test for setting time and Kruskal-Wallis and Dunn test for biocompatibility at 5% significance level. Results: Histologic observation showed no statistically significant difference of biocompatibility (p>0.05) among the materials in the subcutaneous tissues. For the setting time, clinker without calcium sulfate showed the shortest initial and final setting times (6.18 s/21.48 s), followed by clinker with 2% calcium sulfate (9.22 s/25.33 s), clinker with 5% calcium sulfate (10.06 s/42.46 s) and MTA (15.01 s/42.46 s). Conclusions: All the tested materials showed biocompatibility and the calcium sulfate absence shortened the initial and final setting times of the white Portland cement clinker.

Biocompatibilidade e tempo de presa do clinker do cimento Portland cinza, com ou sem sulfato de cálcio

Objetivo: avaliar a biocompatibilidade do clinker do cimento Portland cinza, sem e com 2% e 5% de sulfato de cálcio. Métodos: vinte e quatro ratos receberam implantes no tecido subcutâneo, de tubos de polietileno preenchidos com o pó do clinker do cimento Portland cinza, sem ou com 2% e 5% de sulfato de cálcio. Após 15, 30 e 60 dias da implantação, os animais foram mortos e os espécimes preparados para a análise microscópica.Os tempos de presa de cada material foram avaliados de acordo com as especificações n° C266-08 da ASTM. Os dados foram analisados pelos testes ANOVA e de Tukey, para o tempo de presa; e pelo de Kruskal-Wallise de Dunn para a biocompatibilidade, com nível de significância de 5%. Resultados: histologicamente, não se constatou diferença estatística entre os materiais. Conclusão:o clinker sem sulfato de cálcio apresentou tempo de presa inicial de 5 min. e final de 55 min., seguido pelo clinker com 2% de sulfato de cálcio (8-95 min.) e pelo clinker com 5% de sulfato de cálcio (10-110 min.)

Ultrastructural and immunohistochemical study of the influence of fluoride excess on the development of rat incisor tooth buds

Little information is available on the pathogenesis of fluorosis during the fetal and initial postnatal period. In the present study, female rats received 0 (control), 7 or 100 ppm of sodium fluoride in drinking water, one week before breeding and throughout gestation and nursing periods. The hemimandibles of the offspring were collected at 0, 7 and 14 days of postnatal life (n = 5) and processed for morphological analyses by light and electron microscopy, immunohistochemical analysis for amelogenin and morphometric study of enamel matrix and ameloblasts of incisors. The results showed a decrease in matrix production at the secretory phase at all study periods for the 100 ppm group. In this same group, the secretory ameloblasts showed reduction of enamel matrix secretion, disorganization of mitochondrial crests, large vacuoles at the apical portion of the cytoplasm, retention of intracisternal material and dilatation of some cisterns in the rough endoplasmic reticulum. In the groups of animals aged 7 and 14 days, analysis of variance showed significant reduction (p<0.05) in cytoplasmic volume of 23.80 percent and 24.75 percent, respectively, in relation to the control group. The smooth-ended maturation ameloblasts exhibited a large number of vacuoles with electron-dense endocytic matrix, suggesting a delay in the resorption process. Immunohistochemical analysis showed no difference in the intensity and labeling pattern of the enamel matrix in any study group. Interestingly, in offspring at the age of 14 days for the 7 ppm group, there was an increase in the matrix length at the secretory phase. Therefore, part of the excessive dose of sodium fluoride given to the mother in drinking water can reach the offspring through the placenta and mother's milk, causing morphological changes in ameloblasts and suggesting a reduction in secretion and a delay in matrix resorption.

Healing of root perforations treated with Mineral Trioxide Aggregate (MTA) and Portland cement

Fourteen root perforations were performed for microscopic evaluation of the repair of interradicular tissue in dogs' teeth. These perforations were accomplished at low-speed with a STP 58 bur at the cervical third of the mesial root toward the furcation under irrigation with saline solution, followed by immediate sealing with ProRoot MTA, MTA-Angelus and white Portland cement. The dogs were killed after 90 days, revealing good results. The Kruskal-Wallis test did not demonstrate any statistically significant difference. It was concluded that the three materials showed good sealing in mineralized tissue, with complete closure, and they were free of inflammation in most teeth.

Avaliou-se o reparo de perfurações em dentes de cães, tratadas com ProRoot MTA, MTA Angelus e cimento Portland branco. As perfurações foram feitas na região de furca de premolares, superiores e inferiores, com broca STP 58 sob refrigeração com soro fisiólogico. Os animais foram mortos após 90 dias e os dentes foram preparados para análise microscópica pela coloração da hematoxilina e eosina. Os três materiais propiciaram o selamento da perfuração com tecido mineralizado e o teste de Kruskal-Wallis demostrou não haver diferença estatística entre eles.

Healing of root perforations treated with Mineral Trioxide Aggregate (MTA) and Portland cement%Reparo de perfurações radiculares tratadas com Agregado Trióxido Mineral (MTA) e cimento Portland

Avaliou-se o reparo de perfurações em dentes de cães, tratadas com ProRoot MTA, MTA Angelus e cimento Portland branco. As perfurações foram feitas na região de furca de premolares, superiores e inferiores, com broca STP 58 sob refrigeração com soro fisiólogico. Os animais foram mortos após 90 dias e os dentes foram preparados para análise microscópica pela coloração da hematoxilina e eosina. Os três materiais propiciaram o selamento da perfuração com tecido mineralizado e o teste de Kruskal-Wallis demostrou não haver diferença estatística entre eles.

Efeitos da inalação passiva da fumaça de cigarro sobre as pregas vocais de ratos / Effects of passive smoke inhalation on the vocal cords of rats

Poucos estudos demonstram as reações patológicas devidas à inalação de fumaça pelas vias aéreas de ratos. OBJETIVO: Estudar e analisar os possíveis efeitos histopatologicos produzidos pela inalação crônica de fumaça de cigarro nas pregas vocais de ratos. DESENHO DE ESTUDO: Experimental. MATERIAL E MÉTODOS: 36 ratos masculinos (Rattus norvergicus Wistar), de 60 dias de idade, foram mantidos em gaiolas e expostos à inalação da fumaça produzida por 10 cigarros, 3 vezes ao dia, 7 dias por semana, para períodos de 25, 50 e 75 dias, e os controles respectivos. Os animais foram sacrificados e as suas laringes dissecadas e submetidas a análise histológica com coloração de Hematoxilina e Eosina e analisados através de microscopia simples. RESULTADOS: Os ratos expostos ao cigarro exibiram menor (p <0,05) massa corporal que o grupo controle. Havia hiperplasia e metaplasia escamosa na extremidade livre da prega vocal e hiperplasia escamosa na porção mediana da prega vocal em todos os 3 períodos de estudo. Além disso, o grupo de 50 dias revelou metaplasia com queratinização nesta área. Não foram observadas alterações morfológicas em outras áreas da laringe e reação inflamatória da lâmina propria. CONCLUSÃO: Foi concluído que a inalação passiva de fumaça de cigarro rende mudanças morfológicas importantes no epitélio da prega vocal que podem, eventualmente, progredir para neoplasia.

Allometric analysis of the postnatal growth of rat submandibular gland

The rat submandibular gland grows significantly during the first 10 weeks of postnatal life. During this growth, there is differentiation and maturation of the definitive glandular structures, (acini, intercalated ducts, convoluted granular tubules, striated ducts and excretory ducts) within a highly vascularized stroma. In this study, the absolute volume of each glandular component during postnatal development was determined morphometrically. The increases in gland mass and component volumes were analyzed allometrically relative to the growth of body mass, using Wald’s non-parametric method. The allometric growth of gland mass was monophasic and negative (k<1), with k = 0.86. The absolute volumes of the acini plus terminal tubules, intercalated ducts, striated ducts and excretory ducts all showed a biphasic pattern, with the first phase occuring from day 2 to day 28 and the second phase from day 28 to day 96. In the first phase, all of the structures showed positive allometric growth (k>1), with k values from 1.09, 1.15, 1.49 and 1.17, for the acini plus terminal tubules, intercalated ducts, striated ducts and excretory ducts, respectively, while in the second phase, all showed negative allometrtic growth (k<1), with k values of 0.72, 0.33, 0.77 and 0.82, respectively. The convoluted granular tubules showed a single phase of positive allometric growth (k>1) between 28 and 96 days of age, with k=1.28, whereas the stromal volume showed negative allometric growth (k<1) from day 14 to day 96, with k=0.77.

A stereological analysis of the heterotopic osteogenesis induced by allogenic bone matrix graafts in rat subcutaneous tissue

The subcutaneous cellular and tissue alterations that occur during heterotopic osteogenesis induced by a demineralized bone matrix graft in rats were studied morphometrically. Diaphysis segments obtained from 42 rat femurs were cleaned and demineralized with 0. 6 M HCl. Each segment was filled with a blood clot and implanted into the back of rats (1 segment/rat). Histological speciemens were collected 2, 5, 7, 14, 21, 28 and 49 days after implantation (6 rats/period) and processed after demineralization. Qualitative and morphometric analysis of the histological sections showed that the external blood clots and those filling the medullary canal were replaced with connective tissue within 2-5 days pos-implantation. After 14 days, 11, 2 per cent and 0, 5 per cent of the implanted matrix was replaced with areas of resorption nd hyaline cartilage, respectively. After 21 days, 13, 7 per cent, 2, 4 per cent and 2, 7 per cent of the graft matrix showed areas of resorption, cartilage tissue and bone tissue, respectively. At the end of the 49-day study period, 20, 2 per cent, 7, 8 per cent and 14, 5 per cent of the matrix was replaced with areas of resorption, bone tissue and hematopoietic tissue, respectively. Bone induction in rat subcutaneous tissue occurred only at the site of allogenic matrix implantation. the quantity of neoformed cartilage, bone and myeloid tissue depended on the volume of matrix implanted and on the microenviromental conditions, and was regulated temporally by a cascade of events mediated by bone morphogenetic proteins (BMPs) presents in the graft. These results also show that morphometric methods may be useful for assessing osteoinduction at ectopic sites.

Papillary cystadenoma of the minor salivary gland of the lower lip

A case report of the papillary cystadenoma from minor salivary gland in lower lip of a 54-year-old man is described.

Análise morfométrica e imunohostoquímica da atrofia da glândula parótida do rato provocada pela ligadura do ducto excretor principal / Morphometric and immunohistochemic analysis of rat parotid gland atrophic after principal excretory duct ligation

Os eventos que ocorrem na atrofia da glândula parótida do rato pósligadura cirúrgica do seu dueto principal, foram avaliados por métodos morfométricos e imuno-histoquímicos para proliferação celular (método do PCNA) e para apoptose (método do TUNEL) nos períodos de zero hora e 2, 4, 6 e 8 dias pós-obstrução. A análise dos resultados mostrou que a massa glandular diminuiu cerca de 30% entre os períodos de zero hora e 2 dias, estabilizando-se a seguir. 0 volume total dos ácinos decaiu 87% entre zero hora e 6 dias, devido a redução no volume da célula acinosa e principalmente pela queda linear no número dessas células, com velocidade de decaimento de -32,66 x '10 POT.6' células/dia e maior índice de apoptose no segundo dia. Nesse mesmo período o estroma aumentou 69%, devido inicialmente ao edema e depois ao acréscimo de matriz extracelular, mas também pelo aumento linear no número de células estromais, a uma velocidade de 21,34 x '10 POT.6' células/dia e com maior índice de proliferação no quarto dia. 0 volume total do sistema de ducto aumentou cerca de 18% no período de zero hora e 6 dias, devido a dilatação principalmente dos duetos excretores e ao aumento no número de células. Concluímos, que na atrofia da parótida do rato após ligadura do seu dueto principal, ocorre apoptose e proliferação celular nos seus vários compartimentos morfológicos, com marcante predomínio do primeiro processo no parênquima, especialmente nos ácinos e do segundo no estroma; e que a estabilidade da massa glandular à partir do segundo dia, apesar do marcante declínio no parênquima, foi devido a sua substituição compensatória pelo estroma, provavelmente para preservar o espaço anatômico da parótida

Comparative stereologic study between secretory and maturation ameloblasts in rat incisors.

The ameloblasts both in secretion phase and in smooth-ended ameloblasts in maturation phase were studied using stereologic methods in transmission electron microscopy (TEM). From secretion to maturation phase of amelogenesis, the nucleus volume decreased 23% and cytoplasm volume did not show significant changes; the total volume and surface of the rough endoplasmic reticulum (RER) decreased 74% and 90%, respectively, and of the mitochondria increased 742% and 384%, respectively; the surface-to-volume ratio for RER and mitochondria decrease 59% and 42%, respectively; and the predominantly apical secretory granules disappeared joined at Tomes’ process and lysosomes and phagic vacuoles have appeared principly in supranuclear cytoplasm. Although significant morphologic variation occurs from the secretory to the maturation phase of the ameloblasts, their cytoplasm volume remains unaltered.

Growth of the mouse pancreas during postnatal development

The postnatal growth of the mouse pancreas was studied using stereological methods. The measurements obtained included gland mass, total cell number, the number and frequency of cells in each morphological comportment and the nuclear and cytoplasmic volume of the acinar cells. Pancreatic mass increased significantly (>10,000 fold) in the first 70 days of life and this was accompanied by an increase of 6,841 per cent, 7,027 per cent, 4,864 per cent and 3,360 per cent, respectively. During the same period, the mean acinar cell volume increased by only 146 per cent. These results showed that growth of the mouse pancreas during postnatal development, occurred through intense profiferative activity of all cell types and by an increase in the size of individual cells, notably the acinar cells.

Stereological analysis of the guinea pig pancreas during postnatal development

The postnatal development of the guinea pig pancreas (Cavia porcellus) from 2 to 140 days old was studied stereologically by light microscopy. During this period, the pancreatic volume increased 805 percent. The compartmental volumes of the acini, intercalated ducts, excretory ducts, islets of Langerhans and stroma increased 756 percent, 1,372 percent, 1,591 percent, 3,393 percent and 793 percent, respectively, whereas the total external surfaces of the acini, intercalated ducts and excretory ducts increased 542 percent, 1,667 percent and 1,002 percent, respectively. This growth involved increases of 361 percent, 74 percent, 1,355 percent and 673 percent in the total number of acinar, centroacinar, intercalated duct and excretory duct cells, respectively. In the same period, the mean volume of acinar cells increased 210 percent. These results indicate that the growth of the acinar compartmental volume involves a significant proliferative activity of the acinar cells and an increase in the individual cell volume. Analysis of the surface-to-volume ratio during this growth showed that the acini increased significantly in size. On the other hand, the growth of the parenchymal structures was accompanied by a reduction on interacinar connective tissue spaces and by the final organization of the interlobar and interlobular septa.

A morphometric study of seasonal influences on the daily cycle of rat submandibular glands

The submandibular glands of rats raised at room temperature show daily and seasonal variations. To quantify these variations, the submandivular glands of male Wistar rats were collected at 4h intervals over a 24h period in the summer and winter and processed for histological analysis. Gland weight decreased 37 per cent (P<0.01) in the winter and 13 per cent in the summer. The volume of the acinar compartment decreased 73 per cent (P<0.01) between midnight and 4 a.m. in the winter and 38 per cent (P<0.01) between 8 p.m. and midnight in the summer. In contrast, the volume of the stroma increased about 164 per cent (P<0.01) between midnight and 4 a.m. in the winter and by 10 per cent (P<0.01) between 8 p.m. and midnight in the summer. the volume of the duct system generally remained constant, although in the winter the convoluted granular tubules diminished 44 per cent (P<0.01) between midnight and 4 a.m. Although the volume of the intercalated ducts remained constant over 24h in both seasons, there was a 49 per cent decrease in winter compared to summer (P<0.01). These results indicate that the large changes in gland weight were caused mainly by variation in the volume of the acini, convoluted granular bubules ans stroma. The morphological variations were much more pronounced in winter than in summer and this affected the daily saliva production.