Relationship of clonogenic capacity to plating efficiency and vital dye staining of human periodontal ligament cells: implications for tooth replantation

The survival rate of avulsed permanent teeth following replantation is affected primarily by the duration of the extra-alveolar period and the nature of the storage conditions. These factors are believed to strongly affect the viability of periodontal ligament (PL) cells but in vitro assays of cell viability based on vital dye assays are only weakly correlated with the tooth survival rate after replantation. The aim of the present study was to examine the relative dependence of cell membrane integrity, attachment and clonogenic capacity of human PL cells on the temperature and duration of the extra-alveolar period and the type of storage medium. Twenty-four premolar teeth were extracted for orthodontic reasons from 9 patients 11–18 years of age. Teeth were maintained at 4°C or 23°C for 15, 30, 60 or 120 min in either milk or dry conditions. Cell membrane integrity was determined by BCECF/AM dye inclusion. Plating efficiency was determined by measurement of cell attachment at 3 and 6 h. The clonogenic capacity of progenitor cells was estimated by limiting dilution and colony counts. For all assays teeth stored in milk at 4°C showed the highest percentages of BCECF positive, attached cells with clonogenic capacity. Increased storage time (15–120 min) was associated with a 50% relative reduction of BCECF staining and a 5-fold relative reduction of cell attachment regardless of storage conditions. However, the clonogenic capacity of progenitor cells decreased 25-fold over the same duration of storage. These data demonstrate that in vitro assays of clonogenic capacity are much more sensitive to extra-oral storage time and storage conditions than dye inclusion or cell attachment. We suggest that in comparison with in vitro measures of cell membrane integrity, the clonogenic capacity of PL cells is more closely linked to tooth survival rate, probably reflecting the capacity of PL progenitor cells to recolonize the root surface after replantation.     

In vitro viability, mitogenicity and clonogenic capacity of periodontal ligament cells after storage in four media at room temperature

Abstract – The choice of storage medium for preserving traumatically avulsed teeth is important for the success of future replantation. The objective of this study was to compare the effectiveness of four recommended storage media (Hank's balanced salt solution [HBSS], culture medium, α minimal essential medium [α‐MEM], and ViaSpan) to preserve cultured periodontal ligament fibroblasts (PDLF) at room temperature (22°C). PDLF were obtained from explants of extracted healthy human teeth. Plates with confluent PDLF were soaked in the various media for 2, 8 and 24 h at room temperature. A control group was incubated with culture medium at 37°C. After incubation, viability of the cells was determined by trypan blue exclusion test. Viable cells were then analyzed for mitogenic (with thymidine) and clonogenic capacity (by culturing one cell/well). Viability of PDLF stored up to 24 h was comparable in all tested media, and the differences were limited to 1%–3%. PDLF stored for up to 24 h in various media had statistically comparable mitogenicity to the control group. After 8 h of storage, the differences were limited to 2%–9%, except for the α‐MEM group which had 23%–29% lower mitogenic capacity compared to the control group. Increasing the storage time up to 24 h further decreased the mitogenicity of the cells by 22%–47%. The highest mitogenicity after 24 h of storage was found in PDLF stored in culture medium or HBSS, and the lowest in α‐MEM. PDLF stored for 2–8 h in various media had a comparable clonogenic capacity to the control group. However, after 24 h, the cells' clonogenic ability dropped by 14%–66%. A similar trend of reduction was noted in the mitogenic and clonogenic capacity, although it was statistically significant only in the clonogenic capacity. Culture medium and ViaSpan, followed by HBSS, were the most effective in preserving the clonogenic capacity of PDLF after 24 h of storage. The lowest clonogenic capacity after 24 h of storage was in the α‐MEM group (66%, P<0.0025). In conclusion, culture medium, followed by HBSS and ViaSpan, was the most effective media for preserving the viability, mitogenicity and clonogenic capacity of PDLF stored for up to 24 h at room temperature. The lowest functional abilities were found in PDLF stored in α‐MEM.     

In vitro viability, mitogenicity and clonogenic capacity of periodontal ligament cells after storage in six different media

Abstract— The choice of storage medium for preserving traumatic-ally avulsed teeth is important for the success of future replantation. The objectives of this study were to evaluate the effectiveness of six different media: culture medium, α minimal essential medium (αMEM), milk, Hank's balanced salt solution (HBSS), ViaSpan and conditioned medium (CM) to preserve cultured periodontal ligament fibroblasts (PDLF). Periodontal ligament fibroblasts were obtained from explants of human healthy extracted teeth. Plates with confluent PDLF were soaked in the various media for 2, 8 and 24 h at 4°C. A control group was incubated with culture medium at 37°C. After incubation, cell viability was determined by trypan blue exclusion test. Viable cells were then analyzed for mitogenic (with thymidine) and clonogenic capacity (by culturing one cell/well). Storage of PDLF up to 24 h decreased their vitality by only 2%-14%. Vitality of the PDLF after 2, 8 and 24 h was highest when stored in milk or HBSS (91%-97%) and lowest when stored in ViaSpan or CM (82%-93%). PDLF stored for 2–8 h in various media had a mitogenic capacity comparable to the control. However, increasing the storage period to 24 h decreased the mitogenicity of the cells by 3%-39%. The highest mitogenicity was found in PDLF stored in milk or HBSS and the lowest in CM or ViaSpan. The clonogenic capacity of the cells dropped by 38%-71% after 24 h and was the best indicator of the deteriorating effect of long storage. Milk and HBSS were the most effective in preserving the clonogenic capacity. Nevertheless, reduction in the viability, mitogenicity or clonogenic capacity was statistically significant in nearly all the tested media only after 24 h of incubation. In conclusion, HBSS and milk were the most effective media for preserving the viability, mitogenicity and clonogenic capacity after storage for up to 24 h at 4°C.     

Assessment of post-traumatic PDL cells viability by a novel collagenase assay

Abstract – Both length of extra-alveolar time and type of storage media are significant factors that can affect the long-term prognosis for replanted teeth. Numerous studies have examined various media in an attempt to determine the ideal material for storage of the avulsed tooth. The purpose of this study was to compare the number of viable periodontium ligament (PDL) cells in different storage media using a collagenase assay. Thirty-three freshly extracted human teeth were divided into four experimental and two control groups. The positive and negative controls corresponded to 0 min and an 8-h dry time, respectively. The experimental teeth were stored dry for 30 min and then immersed in one of four media (Hank's balanced salt solution (HBSS), milk, saline, water) for 45 min. The teeth were then treated with dispase grade II and collagenase for 30 min. The number of viable and nonviable PDL cells was counted with a hemocytometer and analyzed. An anova demonstrated no statistically significant differences in the viability of PDL cells among saline, HBSS and milk. Within the parameters of this study, it appears that milk or saline is an equally viable alternative to HBSS for storage of avulsed teeth.     

Tooth replantation after keeping the avulsed tooth in oral environment: case report of a 3-year follow-up

Abstract –  Clinical practice has shown that most avulsed teeth are replanted after an extra-alveolar time that compromises the prognosis of replantation. In cases of delayed replantation, the use of adequate media for storage and transportation of the avulsed teeth may improve this prognosis considerably. Difficulties inherent to accidental dental avulsion include the lack of immediate access to ideal storage media, which accentuates the importance of saliva as a viable and readily available option. The authors report the case of an accidentally avulsed permanent maxillary central incisor that was kept into the patient's oral cavity from the moment of trauma until its replantation, 90 min later. Three years of follow-up revealed absence of root resorption, ankylosis or abnormal mobility, which demonstrates the feasibility of keeping avulsed teeth in saliva, at least when more indicated storage media are not available immediately.     

In vitro viability, mitogenicity and clonogenic capacities of periodontal ligament fibroblasts after storage in four media supplemented with growth factors

Abstract – The choice of storage medium for preserving traumatically avulsed teeth is important for the success of future replantation. The objective of this study was to evaluate the effectiveness of growth factors (IGF‐1 and PDGF‐BB) when added to storage media in preserving the functional abilities of cultured periodontal ligament fibroblasts (PDLF). The evaluated storage media were: ViaSpan, Hanks’ balanced salt solution (HBSS), α minimal essential medium (α MEM), and α MEM supplemented with FCS and antibiotic (α MEM‐S). PDLF were obtained from explants of human healthy extracted teeth. Plates with confluent PDLF were soaked in the various media supplemented with IGF‐1 (10 ng/ml) and PDGF‐BB (4 ng/ml) for 2, 8 and 24 h at room temperature (24 °C). The control group was incubated with the examined storage media without growth factors at 24 °C. An additional control group was incubated with culture medium at 37 °C without growth factors. After incubation, the viability of the cells was determined by Trypan blue exclusion test. Viable cells were then analyzed for mitogenic (with thymidine) and clonogenic (by culturing one cell/well) capacities. Storage of PDLF with growth factors (GF) for 2, 8 and 24 h decreased their vitality by only 3% (not statistically significant). The mitogenicity of PDLF stored for 2, 8 and 24 h in various media with GF was statistically comparable to that of the control group. Generally, the highest mitogenic capacity of PDLF stored with or without GF was found after 8 h of storage. Increasing the storage period to 24 h decreased the mitogenic capacity of the cells stored with GF by only 10–40% compared to the control group. In contrast, the clonogenic capacity of PDLF stored with GF increased with increasing storage periods by 100–300%, and the highest clonogenic capacity was found in most storage media after 24 h of storage with GF. The highest clonogenic and mitogenic capacities were found in cells stored in HBSS followed by α MEM‐S. The mitogenic and clonogenic capacities of PDLF stored in various media supplemented with GF for 2–8 h were generally lower than without GF supplementation. The mitogenic and clonogenic effects of GF‐supplementation was observed only after 24 h of storage. After 24 h of storage with GF, the clonogenic capacity increased by 8–224% and the mitogenicity by 20–37%, except in cells stored in α MEM (‐1%). However, these differences were generally not statistically significant. In conclusion, the mitogenic and clonogenic effects of GF were observed only after 24 h of storage at room temperature. HBSS and α MEM‐S supplemented with GF were the most effective media for preserving the viability, mitogenicity and clonogenic capacity of PDLF stored for 24 h at room temperature. For short periods of storage (2 and 8 h), HBSS and α MEM‐S without GF were preferable.     

In vitro viability, mitogenicity and clonogenic capacity of periodontal ligament cells after storage in six different media

The choice of storage medium for preserving traumatically avulsed teeth is important for the success of future replantation. The objectives of this study were to evaluate the effectiveness of six different media: culture medium, alpha minimal essential medium (alpha-MEM), milk, Hank's balanced salt solution (HBSS), ViaSpan and conditioned medium (CM) to preserve cultured periodontal ligament fibroblasts (PDLF). Periodontal ligament fibroblasts were obtained from explants of human healthy extracted teeth. Plates with confluent PDLF were soaked in the various media for 2, 8 and 24 h at 4 degrees C. A control group was incubated with culture medium at 37 degrees C. After incubation, cell viability was determined by trypan blue exclusion test. Viable cells were then analyzed for mitogenic (with thymidine) and clonogenic capacity (by culturing one cell/well). Storage of PDLF up to 24 h decreased their vitality by only 2%-14%. Vitality of the PDLF after 2, 8 and 24 h was highest when stored in milk or HBSS (91%-97%) and lowest when stored in ViaSpan or CM (82%-93%). PDLF stored for 2-8 h in various media had a mitogenic capacity comparable to the control. However, increasing the storage period to 24 h decreased the mitogenicity of the cells by 3%-39%. The highest mitogenicity was found in PDLF stored in milk or HBSS and the lowest in CM or ViaSpan. The clonogenic capacity of the cells dropped by 38%-71% after 24 h and was the best indicator of the deteriorating effect of long storage. Milk and HBSS were the most effective in preserving the clonogenic capacity. Nevertheless, reduction in the viability, mitogenicity or clonogenic capacity was statistically significant in nearly all the tested media only after 24 h of incubation. In conclusion, HBSS and milk were the most effective media for preserving the viability, mitogenicity and clonogenic capacity after storage for up to 24 h at 4 degrees C.     

A quantitative analysis of Propolis: a promising new storage media following avulsion

Abstract –  Both length of extra-alveolar time and type of storage media are significant factors that can affect the long-term prognosis of replanted teeth. Numerous studies have examined various media in an attempt to determine the ideal material for storage of the avulsed tooth. The purpose of this study was to use a Collagenase–Dispase assay to investigate the potential of a new storage media, Propolis, in maintaining viable periodontal ligament (PDL) cells on simulated avulsed teeth. Seventy freshly extracted human teeth were divided into five experimental groups and two control groups. The positive and negative controls corresponded to 0-min and an 8-h dry time, respectively. The experimental teeth were stored dry for 30 min and then immersed in one of the five media (Hank's balanced salt solution (HBSS), milk, saline, Propolis 50%, and Propolis 100% for 45 min). The teeth were then treated with dispase grade II and collagenase for 30 min. The number of viable PDL cells were counted with a hemocytometer and analyzed. Statistical analysis demonstrated that both Propolis groups kept significantly more PDL cells viable compared to either milk, saline, or HBSS. Within the parameters of this study, it appears that Propolis may be a better alternative to HBSS, milk, or saline in terms of maintaining PDL cell viability after avulsion and storage.     

Evaluation of the topical effect of alendronate on the root surface of extracted and replanted teeth. Microscopic analysis on rats' teeth

Abstract –  The treatment of choice for tooth avulsion is replantation. The ideal replantation should be realized as quickly as possible, or at least, the avulsed tooth should be kept in an adequate solution to preserve the periodontal ligament attached to the root. If that is not possible, treatment of the radicular surface should be done in order to prevent radicular resorption. The purpose of this study was to test sodium alendronate as a substance for topical treatment of the radicular surface of avulsed teeth in an attempt to prevent the occurrence of dental resorptions. Fifty-four rat maxillary right central incisors were extracted and replanted. Group I – extra-alveolar dry period of 15 min, intracanal dressing with calcium hydroxide (CALEN^®, S.S. White, Artigos Dentários LTDA, Rio de Janeiro, Brazil) and replantation; Groups II and III – extra-alveolar dry periods of 30 and 60 min, respectively, immersion in 1% sodium hypochlorite for 30 min for removal of the periodontal ligament, washing in saline solution for 5 min, and treatment of the radicular surface with 3.2 mg/l sodium alendronate solution for 10 min. Intracanal dressing with calcium hydroxide and replantation followed. At 15, 60, and 90 days post-reimplantation, the animals were killed and the samples obtained and processed for microscopic analysis. The results indicated that sodium alendronate was able to reduce the incidence of radicular resorption, but not of dental ankylosis. No significant differences were observed regarding variations in the extra-alveolar periods among the groups.     

Effect of extra-alveolar period and storage media upon periodontal and pulpal healing after replantation of mature permanent incisors in monkeys

The effect of extra-alveolar period and storage media upon periodontal and pulpal healing after replantation was studied in green Vervet monkeys (Cercopithecus aethiops). Mandibular lateral incisors were extracted. The extra-alveolar period before replantation was 0, 18, 30, 60, 90, and 120 min. The storage media for the extracted teeth were tap water, physiologic saline, saliva or dry storage. The animals were sacrificed 8 weeks after replantation and the replanted teeth were examined histometrically. The following histologic parameters were registered for each tooth: surface resorption, inflammatory resorption, replacement resorption (ankylosis), periapical inflammatory changes, the extent of vital pulp and downgrowth of pocket epithelium. A significant relationship was found between the frequency of root resorption, extra-alveolar period and storage medium. This was especially evident after dry storage. Surface resorption was found with approximately the same frequency irrespective of extra-alveolar period and storage media. Inflammatory root resorption was especially common after dry storage and was related to the length of the extra-alveolar period. Already after 30 min dry storage, this resorption type was very prominent. Teeth stored in tap water, saline or saliva showed about the same frequency of inflammatory resorption, which increased slightly with increased extra-alveolar periods. Replacement resorption showed a strong relationship to dry storage and became very prominent after 60 min. Replacement resorption was rarely found among teeth stored in saline or saliva; whereas it was significantly increased among teeth stored in tap water. It is concluded that saline and saliva offer good protection against root resorption during the extra-alveolar period.     

Viability of fibroblasts in a novel probiotic storage media

Abstract – A number of storage media have been investigated as to their ability to maintain the viability of the periodontal ligament (PDL) cells and thus to permit longer extra‐alveolar periods prior to replantation of avulsed teeth. The aim of the present in vitro study was to evaluate the number of viable PDL cells of avulsed teeth treated by Hank’s Balanced Salt Solutions (HBSS), saline, a novel probiotic solution and milk. Thirty‐six freshly extracted single‐rooted human teeth with closed apices were divided into one of the four experimental groups and two control groups (N = 6 each). The positive and negative controls corresponded to 0 min and an 8‐h dry time respectively. Following extraction, the coronal 3 mm of PDL tissue was scraped with a #15 scalpel to remove cells that might have been damaged. The experimental teeth were dried for 30 min followed by a 45 min immersion in one of the four experimental media. Each experimental tooth, after drying and soaking, was incubated for 30 min with a 2.5 ml solution of 0.2 mg ml^?1 of collagenase CLS II and a 2.4 mg ml^?1 solution of dispase grade II in phosphate buffer saline (PBS). The cells were then labelled with 0.4% Trypan blue for determination of viability. The teeth stored in positive control demonstrated the highest number of viable PDL cells followed in rank order by HBSS, saline, Lactobacillus reuteri solution and milk. There was no significant difference in the number of viable PDL cells between HBSS, milk, L. reuteri solution and saline. Within the parameters of this study, it appears that probiotic may be able to maintain PDL cell viability as HBSS, milk, or saline.     

Pedialyte Promotes Periodontal Ligament Cell Survival and Motility

IntroductionThe search still continues to find the best storage media for avulsed teeth. Unfortunately, some of the recommended storage solutions are not commonly found in households or do not preserve the periodontal ligament (PDL) cells long-term. The purpose of the present study was to determine whether Pedialyte is a viable alternative storage solution for avulsed teeth by assessing its ability to preserve human PDL cell viability.MethodsHuman PDL cells were exposed to 6 different storage solutions (minimal essential medium [MEMα], Hank's balanced salt solution [HBSS], non-fat milk, coconut water, Pedialyte, or tap water) for 2, 6, 24, or 48 hours at 4°C or 25°C. Cell viability was quantified immediately or 1 week after exposure. The effects of these storage solutions on PDL cell motility and bacterial proliferation were also examined. The results were statistically analyzed by analysis of variance.ResultsPedialyte at 4°C and 25°C showed significantly (P < .001) higher cell survival compared with water after all time intervals. No significant difference was noted between control (MEMα), HBSS, coconut water, and Pedialyte at 4°C after 2 hours. Cells stored in Pedialyte for 24 hours at 25°C and assayed 1 week later showed significantly higher cell survivability compared with milk. Pedialyte supported significantly less bacterial growth compared with non-fat milk and coconut water. No difference in cell motility was observed for cells stored for 24 hours in Pedialyte, MEMα, HBSS, milk, or coconut water.ConclusionsPedialyte is a viable alternative as a storage solution for avulsed teeth.     

不同离体时间和保存液对犬离体牙牙周膜细胞活力影响的实验研究

目的:探讨完全脱位牙不同离体时间和保存液对牙周膜细胞活力的影响。方法:麻醉拔除犬牙35个,首先将20个牙随机分为5组,分别为室温干燥放置0、30、60、120、240 min组,另15个牙室温干燥放置30 min后,随机分为3组,分别放入牛奶、HBSS液,100 g/L蜂胶液中浸泡2 h。各组处理完成后,采用全牙消化法获得牙周膜细胞,并通过4 g/L台盼蓝染色法检测各组牙周膜活细胞数和存活率。结果:室温干燥放置30、60、120、240 min后,牙周膜细胞存活率依次为33.6%、23.6%、18.5%、0.8%,而0 min的牙周膜细胞存活率可达95.5%。拔后30 min,经牛奶、HBSS液和100 g/L蜂胶液中保存2 h后,牙周膜细胞均有活力,其细胞存活率大小依次为100 g/L蜂胶液、HBSS液和牛奶,其中100 g/L蜂胶液与HBSS液相比无统计学差异(P>0.05),但与牛奶组相比,均有统计学差异(P<0.05)。结论:随着离体时间延长,完全脱位牙根面牙周膜细胞活力明显下降。100 g/L蜂胶液和HBSS液保存犬牙牙周膜细胞活力优于牛奶液。     

In vitro viability, mitogenicity and clonogenic capacity of periodontal ligament cells after storage in four media at room temperature

The choice of storage medium for preserving traumatically avulsed teeth is important for the success of future replantation. The objective of this study was to compare the effectiveness of four recommended storage media (Hank's balanced salt solution [HBSS], culture medium, alpha minimal essential medium [alpha-MEM], and ViaSpan) to preserve cultured periodontal ligament fibroblasts (PDLF) at room temperature (22 degrees C). PDLF were obtained from explants of extracted healthy human teeth. Plates with confluent PDLF were soaked in the various media for 2, 8 and 24 h at room temperature. A control group was incubated with culture medium at 37 degrees C. After incubation, viability of the cells was determined by trypan blue exclusion test. Viable cells were then analyzed for mitogenic (with thymidine) and clonogenic capacity (by culturing one cell/well). Viability of PDLF stored up to 24 h was comparable in all tested media, and the differences were limited to 1%-3%. PDLF stored for up to 24 h in various media had statistically comparable mitogenicity to the control group. After 8 h of storage, the differences were limited to 2%-9%, except for the alpha-MEM group which had 23%-29% lower mitogenic capacity compared to the control group. Increasing the storage time up to 24 h further decreased the mitogenicity of the cells by 22%-47%. The highest mitogenicity after 24 h of storage was found in PDLF stored in culture medium or HBSS, and the lowest in alpha-MEM. PDLF stored for 2-8 h in various media had a comparable clonogenic capacity to the control group. However, after 24 h, the cells' clonogenic ability dropped by 14%-66%. A similar trend of reduction was noted in the mitogenic and clonogenic capacity, although it was statistically significant only in the clonogenic capacity. Culture medium and ViaSpan, followed by HBSS, were the most effective in preserving the clonogenic capacity of PDLF after 24 h of storage. The lowest clonogenic capacity after 24 h of storage was in the alpha-MEM group (66%, P < 0.0025). In conclusion, culture medium, followed by HBSS and ViaSpan, was the most effective media for preserving the viability, mitogenicity and clonogenic capacity of PDLF stored for up to 24 h at room temperature. The lowest functional abilities were found in PDLF stored in alpha-MEM.     

Evaluation,using extracted human teeth,of Ricetral as a storage medium for avulsions – an in vitro study

Abstract – The prognosis of teeth replanted following avulsion is determined by the extra‐alveolar time and storage medium used. This study was undertaken to determine the efficacy of an oral rehydration solution ‘Ricetral’, in retaining the vitality of periodontal ligament cells when used as a storage medium for avulsed teeth prior to replantation. The study consisted of a comparative evaluation between Ricetral and two currently recommended solutions, Hank’s balanced salt solution (HBSS) and milk. Thirty extracted teeth were dried for 30 min and soaked in the respective storage media for 45 min. The periodontal ligament cells were isolated by an enzyme treatment with collagenase and trypsin. The cells were evaluated for vitality by trypan blue staining and number of vital cells counted in a hemocytometer. Statistical analysis revealed that cell vitality was high with Ricetral and HBSS, but poor with milk.     

Current developments in interim transport (storage) media in dentistry: an update

Healing following avulsion and replantation is dependent on the extent of pulpal and periodontal ligament (PDL) tissue damage. Therefore, immediate replantation is the recommended treatment of choice for an avulsed permanent tooth. To achieve a more favourable prognosis following tooth replantation, use of an appropriate interim transport medium is usually advocated. Numerous studies have researched and advocated the use of media like saliva, milk, Hank's Balanced Salt Solution (HBSS) and ViaSpan. However, current research has indicated the use of few newer media as promising interim transport media for an avulsed tooth. This review summarises the current developments regarding the introduction of newer interim transport media for the treatment of avulsed teeth.     

Delayed replantation of rat teeth after use of reconstituted powdered milk as a storage medium

Abstract – Minimal extraoral dry storage period and moist storage for the avulsed tooth are identified as key steps for the treatment protocol of tooth replantation. Among the possible moist storage media, bovine milk has stood out because of its capacity of preserving the integrity of the periodontal ligament (PDL) fibers. This condition has attracted the attention to investigate the use of powdered milk, which is one of the presentation forms of bovine milk, as a feasible storage medium in cases of delayed tooth replantation. The aim of this study was to evaluate the healing process after delayed replantation of rat teeth stored in reconstituted powdered milk and long shelf‐life (ultra high temperature) whole milk. Forty maxillary right rat incisors were assigned to four groups (n = 10): group I – the teeth were extracted and immediately replanted into theirs sockets; group II – the teeth were stored for 60 min in 200 ml of freshly reconstituted powdered milk; group III – the teeth were stored for 60 min in 200 ml of long shelf‐life whole milk; group IV – the teeth were kept dry for the same time. All procedures were performed at room temperature. Next, the root canals of teeth in groups II, III, and IV were instrumented, filled with a calcium hydroxide‐based paste, and replanted into their sockets. All animals received systemic antibiotic therapy and were killed by anesthetic overdose 60 days after replantation. The pieces containing the replanted teeth were removed, fixed, decalcified, and paraffin‐embedded. Semi‐serial 6‐μm‐thick sections were obtained and stained with hematoxylin and eosin for histomorphological analysis. There was statistically significant difference (P < 0.05) between groups I and IV regarding the presence of replacement resorption and PDL remnants on root surface. The powdered milk and long shelf‐life whole milk presented similar results to each other and may be indicated as storage media for avulsed teeth.     

Investigation of the children referred to a dental hospital with avulsed permanent incisor teeth

Abstract The management of children referred to a dental hospital because of avulsed permanent incisor teeth was considered in a retrospective study. The group consisted of 49 children between the ages of 6 and 14 years. Most of the children (60%) had their avulsed teeth stored dry while only 7% had them stored in milk, and 19% in saliva. Following avulsion the majority of the children attended a dental surgeon or general hospital. Twenty-three children contacted health care personnel within 30 minutes but only 13 had their teeth replanted in this time period. Thirty-six children had 46 incisors replanted. No relationship was demonstrated between the place or personnel who replanted the teeth, and a successful outcome.     

Effect of preservation media on proliferation and collagen biosynthesis of periodontal ligament fibroblasts

Abstract The viability of periodontal ligament (PDL) cells, which are responsible for the production of ligament proteins, is essential for clinical healing of replanted exarticulated teeth. Cultured human PDL fibroblast-like cells were used to study the effects of various transport media on cell proliferation, collagen synthesis and total protein synthesis. The PDL cells were obtained from intact premolars of a healthy young male patient by trypsinization and scraping the PDL tissue. Proliferation of the cells, the activity of prolyl 4-hydroxylase, total protein and collagen synthesis were studied after 60 min incubation in Dulbecco's Modified Eagle's Medium (DMEM), milk, saliva and tap water. Proliferation capacity was also investigated after prolonged incubation in milk. The results indicated that milk and saliva were superior to water in maintaining these cell functions, but as the saliva used in this study does not resemble that in the clinical situation, we recommend milk as a temporary storage medium for exarticulated teeth before replantation.     

Mitoses and microorganisms in the periodontal membrane after storage in milk or saliva

Abstract – Milk and saliva were tested in vitro as potential storage media for avulsed teeth. Developing monkey teeth were extracted and stored in milk or saliva for periods ranging from l to 6 h. The osmolality, pH, conductivity and number of viable bacteria in the media were determined after predetermined intervals during the storage periods. After the storage periods the teeth were either prepared for scanning electron microscopy or cultured for 24 h in Eagle's medium supplemented with ³H-thymidine. In the scanning electron microscope numerous adherent bacteria were seen covering the periodontal membrane after storage in saliva but none were found after storage in milk. The cultured teeth were sectioned and evaluated with autoradiography. Superficial parts of the periodontal membrane were rapidly injured by storage in saliva while the epithelial root sheath and the apical pulpal cells were affected at a later stage. Cells neighboring the cementoblasts incorporated ³H-thymidine after 6 h storage in milk but not after storage in saliva for the same length of time. It was concluded that the low osmolality in combination with bacteria which adhered to the periodontal membrane made saliva less suited than milk for long time storage of avulsed teeth. Furthermore, a viable layer of cells close to the root surface seemed to be a prerequisite for a successful healing without root résorption after replantation.