Carsten Staszyk

Equine odontoclastic tooth resorption and hypercementosis

A poorly described, painful disorder of incisor and canine teeth, variably causing periodontitis, with resorptive or proliferative changes of the calcified dental tissues, has recently been documented in aged horses. No plausible aetiopathogenesis for this syndrome has been recorded. Eighteen diseased teeth from eight horses were examined grossly and microscopically and showed the presence of odontoclastic cells by tartrate resistant acid phosphatase (TRAP) staining. A chronological sequence of odontoclastic resorption followed by hypercementosis was demonstrated and, consequently, the term equine odontoclastic tooth resorption and hypercementosis (EOTRH) is proposed for this disorder. EOTRH shares many features with similar dental syndromes described in humans and cats. An aetiological hypothesis proposes mechanical stress of the periodontal ligament as the initiating factor.

Isolation and characterization of multipotent mesenchymal stromal cells from the gingiva and the periodontal ligament of the horse

BackgroundThe equine periodontium provides tooth support and lifelong tooth eruption on a remarkable scale. These functions require continuous tissue remodeling. It is assumed that multipotent mesenchymal stromal cells (MSC) reside in the periodontal ligament (PDL) and play a crucial role in regulating physiological periodontal tissue regeneration. The aim of this study was to isolate and characterize equine periodontal MSC.Tissue samples were obtained from four healthy horses. Primary cell populations were har-vested and cultured from the gingiva, from three horizontal levels of the PDL (apical, midtooth and subgingival) and for comparison purposes from the subcutis (masseteric region). Colony-forming cells were grown on uncoated culture dishes and typical in vitro characteristics of non-human MSC, i.e. self-renewal capacity, population doubling time, expression of stemness markers and trilineage differentiation were analyzed.ResultsColony-forming cell populations from all locations showed expression of the stemness markers CD90 and CD105. In vitro self-renewal capacity was demonstrated by colony-forming unit fibroblast (CFU-F) assays. CFU-efficiency was highest in cell populations from the apical and from the mid-tooth PDL. Population doubling time was highest in subcutaneous cells. All investigated cell populations possessed trilineage differentiation potential into osteogenic, adipogenic and chondrogenic lineages.ConclusionsDue to the demonstrated in vitro characteristics cells were referred to as equine subcutaneous MSC (eSc-MSC), equine gingival MSC (eG-MSC) and equine periodontal MSC (eP-MSC). According to different PDL levels, eP-MSC were further specified as eP-MSC from the apical PDL (eP-MSCap), eP-MSC from the mid-tooth PDL (eP-MSCm) and eP-MSC from the subgingival PDL (eP-MSCsg). Considering current concepts of cell-based regenerative therapies in horses, eP-MSC might be promising candidates for future clinical applications in equine orthopedic and periodontal diseases.

Biomechanical evaluation of the equine masticatory action: Calculation of the masticatory forces occurring on the cheek tooth battery

The forces generated during equine mastication influence the teeth, as well as the periodontium, the jaw bones, the temporomandibular joints and materials used in dental therapy. Due to the limited accessibility of the cheek teeth located farther caudal in the equine oral cavity the measurement of the masticatory forces on these teeth is virtually impossible. The calculation of such forces is an appropriate, indirect alternative to direct measuring. Hence, a mathematical model, which respects several morphological features of the equine dentition, e.g. the Curve of Spee, is presented. The relevant parameters were measured on laterolateral radiographs of 24 cadaver heads and 6 skulls. Our calculations yielded two major results. (1) The masticatory forces increased from rostral to caudal. This increase was due to distinct anatomical features of the equine dentition, i.e. the Curve of Spee, the position of the teeth, and the length of the mandible. (2) The masticatory forces decreased with age. This decrease was due to the reduced height of the Curve of Spee in older horses. These results are discussed in view of veterinary dental treatment, with special reference to the geriatric horse.

Isolation of equine multipotent mesenchymal stromal cells by enzymatic tissue digestion or explant technique: comparison of cellular properties

BackgroundThe treatment of tendon lesions with multipotent mesenchymal stromal cells (MSCs) is widely used in equine medicine. Cell sources of MSCs include bone marrow, as well as solid tissues such as adipose tissue. MSCs can be isolated from these solid tissues either by enzymatic digestion or by explant technique. However, the different preparation techniques may potentially influence the properties of the isolated MSCs. Therefore, the aim of this study was to investigate and compare the effects of these two different methods used to isolate MSCs from solid tissues.Equine adipose tissue, tendon and umbilical cord matrix served as solid tissue sources of MSCs with different stiffness and density. Subsequent to tissue harvest, MSCs were isolated either by enzymatic digestion with collagenase or by explant technique. Cell yield, growth, differentiation potential and tendon marker expression were analysed.ResultsAt first passage, the MSC yield was significantly higher in enzymatically digested tissue samples than in explanted tissue samples, despite a shorter period of time in primary culture. Further analysis of cell proliferation, migration and differentiation revealed no significant differences between MSCs isolated by enzymatic digestion and MSCs isolated by explant technique. Interestingly, analysis of gene expression of tendon markers revealed a significantly higher expression level of scleraxis in MSCs isolated by enzymatic digestion.ConclusionsBoth isolation techniques are feasible methods for successful isolation of MSCs from solid tissues, with no major effects on cellular proliferation, migration or differentiation characteristics. However, higher MSC yields were achieved in a shorter period of time by collagenase digestion, which is advantageous for the therapeutic use of MSCs. Moreover, based on the higher level of expression of scleraxis in MSCs isolated by enzymatic digestion, these cells might be a better choice when attempting tendon regeneration.

Collagen fiber architecture of the periodontal ligament in equine cheek teeth.

The objective of this study was to examine the spatial arrangement of the fiber apparatus of the equine periodontium which is supposed to meet two contrary requirements: (1) to attach the tooth firmly and elastically to the alveolar bone; and, to be appropriately remodeled and reconstructed in order to facilitate the prolonged eruption of the tooth. Specimens of periodontal ligament were obtained from the buccal and lingual/palatal aspects of the first molars from the maxilla and mandible of 12 horses. The animals were assigned to three age groups. Histological sections were prepared from three specific horizontal levels of the periodontal ligament and examined with conventional and polarized-light microscopy. At the gingival level, collagen fascicles (diameter > 200 μm) were densely packed. Their spatial alignment was the same in all age groups. The architecture of the collagen fiber apparatus differed at the middle and apical levels in the three age groups. There was a clear distinction between fibers, bundles, and fascicles. Bundling of collagen fibers, density of the fiber arrangement, and collagen fascicles with an alveolo-cemental orientation increased with age. The collagen fiber apparatus of the equine periodontal ligament is highly adaptive, responding continuously to the dynamic changes in the periodontal environment. Site-specific arrangements and age-dependent structural variations are assumed to maintain tooth support as the reserve crown gradually decreases in length with progressive dental attrition. Most of the age-dependent changes to the periodontal ligament in teeth examined in this study occurred at the apical level. The apical region of young teeth had no periodontal attachment, while the roots of older teeth were firmly attached to the alveolus. When evaluating periodontal ligament development, the individual tooths ‘dental age’ should be considered rather than the animals age to account for individual tooth eruption times.

Influence of mechanical manipulations on the local inflammatory reaction in the equine colon

REASONS FOR PERFORMING STUDY Large intestinal diseases in horses are characterised by inflammation, which could arise from the disease process with some contribution from intestinal manipulation. The effects of the latter are unknown but important to surgeons and could contribute to post operative complications. OBJECTIVES To characterise type and degree of intestinal inflammation induced by various mechanical stimuli in the equine ascending colon. METHODS Laparotomy was performed in 12 horses, the left dorsal colon exteriorised and 3 segments randomly exposed to different mechanical manipulations: 1) enterotomy, 2) enterotomy and mucosal irritation and 3) serosal irritation. Intestinal biopsies were harvested before, immediately after and 30 min after each manipulation for histological evaluation. Eosinophils were detected with Lunas stain and neutrophils identified by immunohistochemical staining for calprotectin. Additionally, left dorsal colon samples from 14 horses from a jejunal ischaemia-reperfusion study were collected immediately after laparotomy (7 horses) and at the end of the experiment without previous manipulation of the colon (7 horses). Horses were subjected to euthanasia at the end of both studies. RESULTS Redistribution of mucosal neutrophils and eosinophils towards the luminal surface and increased neutrophilic infiltration of the submucosa were demonstrated after serosal and mucosal irritation. All manipulations resulted in serosal infiltration with neutrophils. Laparotomy and small intestinal manipulation increased mucosal eosinophilic infiltration. CONCLUSIONS AND POTENTIAL RELEVANCE Mechanical intestinal manipulation caused a rapid local inflammatory reaction in the mucosa, submucosa and serosa including a mucosal eosinophilic response. These changes could exacerbate existing inflammation in horses with large colon disease. Colic surgery can lead to intestinal inflammation in nonmanipulated intestine and this could contribute to a higher morbidity rate in horses after prolonged colic surgery. An intestinal biopsy should be collected at the beginning of surgery to avoid false interpretations.

The dental cavities of equine cheek teeth: three-dimensional reconstructions based on high resolution micro-computed tomography

BackgroundRecent studies reported on the very complex morphology of the pulp system in equine cheek teeth. The continuous production of secondary dentine leads to distinct age-related changes of the endodontic cavity. Detailed anatomical knowledge of the dental cavities in all ages is required to explain the aetiopathology of typical equine endodontic diseases. Furthermore, data on mandibular and maxillary pulp systems is in high demand to provide a basis for the development of endodontic therapies. However, until now examination of the pulp cavity has been based on either sectioned teeth or clinical computed tomography. More precise results were expected by using micro-computed tomography with a resolution of about 0.1 mm and three-dimensional reconstructions based on previous greyscale analyses and histological verification. The aim of the present study was to describe the physiological configurations of the pulp system within a wide spectrum of tooth ages.ResultsMaxillary teeth: All morphological constituents of the endodontic cavity were present in teeth between 4 and 16 years: Triadan 06s displayed six pulp horns and five root canals, Triadan 07-10s five pulp horns and four root canals and Triadan 11s seven pulp horns and four to six root canals. A common pulp chamber was most frequent in teeth ≤5 years, but was found even in a tooth of 9 years. A large variety of pulp configurations was observed within 2.5 and 16 years post eruption, but most commonly a separation into mesial and distal pulp compartments was seen. Maxillary cheek teeth showed up to four separate pulp compartments but the frequency of two, three and four pulp compartments was not related to tooth age (P > 0.05). In Triadan 06s, pulp horn 6 was always connected to pulp horns 1 and 3 and root canal I. In Triadan 11s, pulp horns 7 and 8 were present in variable constitutions. Mandibular teeth: A common pulp chamber was present in teeth up to 15 years, but most commonly seen in teeth ≤5 years. A segmented pulp system was found in 72% of the investigated teeth. Segmentation into separate mesial and distal pulp compartments was most commonly present. Pulp horn 4 coalesced either with the mesial pulp horns 1 and 3 or with the distal pulp horns 2 and 5.ConclusionsDetails of the pulpar anatomy of equine cheek teeth are provided, supporting the continuous advancement in endodontic therapy. Numerous individual configurations of the pulp system were obtained in maxillary cheek teeth, but much less variability was seen in mandibular cheek teeth.

Absence of lymphatic vessels in the dog dental pulp: an immunohistochemical study

In spite of numerous investigations it has not been precisely determined whether lymphatic vessels are present in the dental pulp of dogs. Therefore, this study attempted a specific immunohistochemical detection of lymphatic endothelium. The canine teeth of 19 healthy beagle dogs were dissected into three segments (apical, intermediate and occlusal). After decalcification, specimens were embedded in paraffin wax and histologic cross‐sections were stained immunohistochemically using a reliable antibody (anti‐Prox‐1) against the homeobox transcription factor Prox‐1, which is located within the nucleus of lymphatic endothelium. Anti‐Prox‐1 reacted positively with canine control tissues (lymph nodes, gingiva, nasal mucosa), but showed no staining in tissue sections of the dental pulp. The dog dental pulp contained no vascular structures lined with lymphatic endothelium. This suggests that drainage of interstitial fluid makes use of other routes, i.e. extravascular pathways.

Infundibula of equine maxillary cheek teeth: Part 2: Morphological variations and pathological changes.

Incomplete cemental filling of the infundibula of equine maxillary cheek teeth (CT) is a common feature. Depending on the extent of the defect, three stages of infundibular decay have been suggested. However, histomorphological criteria to identify non-pathological abnormalities and destructive changes have not been defined. Six hundred and eighty eight CT with no evidence of dental diseases and 55 diseased permanent, fully erupted maxillary CT were evaluated on a macroscopic level by assessing the occlusal surface and horizontal sections, including porphyrin assays to detect residual blood within the infundibular cementum. Selected specimens were investigated on a microscopic level using routine and immunohistological staining methods to identify possible routes for the spread of infectious agents from the infundibulum into the endodontic system. Infundibular cemental hypoplasia was defined as a non-pathological developmental abnormality and was detected in >50% of CT with no evidence of dental diseases and in >70% of diseased CT. The first molar (Triadan 09) showed the highest prevalence (75%) of infundibular cemental hypoplasia. The mesial infundibulum was more often affected than the distal infundibulum. Infundibular erosion was considered as the most appropriate term to describe destructive infundibular changes. Infundibular erosion was present in <6% of CT with no evidence of dental diseases, but was detected in >27% of diseased teeth, always accompanied by endodontic disease. This suggests that teeth affected by infundibular cemental hypoplasia are prone to destructive erosion, which possibly leads to endodontic disease. Morphological factors that supplement this ethological hypothesis were described. In 74% of infundibula residual blood was identified, although no vital blood vessels were detected. It is assumed that this content of blood remained in the ample infundibular cemental blood system after tooth eruption and creates a favorable environment for microbial growth. The infundibular enamel was characterised by numerous infoldings to variable degrees and depths. In certain areas focal enamel aplasia was observed. These morphological features might contribute to microbiological settlement and spread of infectious agents through the infundibular wall into the endodontic system.

The sinonasal communication in the horse: examinations using computerized three-dimensional reformatted renderings of computed-tomography datasets

BackgroundSinusitis is a common disease in the horse. In human medicine it is described, that obstruction of the sinonasal communication plays a major role in the development of sinusitis. To get spatial sense of the equine specific communication ways between the nasal cavity and the paranasal sinuses, heads of 19 horses, aged 2 to 26 years, were analyzed using three-dimensional (3D) reformatted renderings of CT-datasets. Three-dimensional models were generated following manual and semi-automated segmentation. Before segmentation, the two-dimensional (2D) CT-images were verified against corresponding frozen sections of cadaveric heads.ResultsThree-dimensional analysis of the paranasal sinuses showed the bilateral existence of seven sinus compartments: rostral maxillary sinus, ventral conchal sinus, caudal maxillary sinus, dorsal conchal sinus, frontal sinus, sphenopalatine sinus and middle conchal sinus. The maxillary septum divides these seven compartments into two sinus systems: a rostral paranasal sinus system composed of the rostral maxillary sinus and the ventral conchal sinus and a caudal paranasal sinus system which comprises all other sinuses. The generated 3D models revealed a typically configuration of the sinonasal communication ways. The sinonasal communication started within the middle nasal meatus at the nasomaxillary aperture (Apertura nasomaxillaris), which opens in a common sinonasal channel (Canalis sinunasalis communis). This common sinonasal channel ramifies into a rostral sinonasal channel (Canalis sinunasalis rostralis) and a caudo-lateral sinonasal channel (Canalis sinunasalis caudalis). The rostral sinonasal channel ventilated the rostral paranasal sinus system, the caudo-lateral sinonasal channel opened into the caudal paranasal sinus system. The rostral sinonasal channel was connected to the rostral paranasal sinuses in various ways. Whereas, the caudal channel showed less anatomical variations and was in all cases connected to the caudal maxillary sinus. Volumetric measurements of the sinonasal channels showed no statistically significant differences (P < 0.05) between the right and left side of the head.ConclusionsUnder physiologic conditions both paranasal sinus systems are connected to the nasal cavity by equine specific sinonasal channels. To resolve sinus disease it is aimed to maintain or even reconstruct the normal anatomy of the sinonasal communication by surgical intervention. Therefore, the presented 3D analyses may provide a useful basis.