Necrotic Layer Research Articles

Obstructive component analysis of radioactive stents and common plastic stents in the bile duct.

Endoscopic placement of a iodine-125 radioactive stent is useful to treat obstructive jaundice with unresectable periampullary tumors. This study aimed to retrospectively evaluate the obstructive component of biliary radioactive stents and discuss the different obstructive mechanism with common plastic stents. Twenty consecutive patients with malignant obstruction underwent insertion of stents into the common bile duct, including 10 radioactive stents and 10 polyethylene stents. The radioactive stents were withdrawn after ∼3 months or earlier if clinical signs suggested stent clogging. Polyethylene stents were withdrawn after physical signs suggested stent clogging. Bacteriologic analyses included identification of aerobic and anaerobic bacteria. Stent surfaces were observed by scanning electron microscopy. Stent deposition was identified by Fourier-transformed infrared spectroscopy and pyrolysis derivatization/gas chromatography/mass spectrometry. Radioactive stent group and polyethylene stent group stents were placed for 86 days (interquartile range 62, 114) and 146 days (interquartile range 105, 181) respectively. The placement duration of the two types of stents was statistically significant. A variety of microorganisms were cultured from the stent deposits. Scanning electron microscope images showed a thicker necrotic layer on the external surface of polyethylene stent than the radioactive stent group. The proportions of obstructive components in each stent were different, but none of them were statistically significant. Necrotic tumor tissue was found in the radioactive stent group. Similar clogging events occurred in both radioactive stents and polyethylene stents. The median duration time of the radioactive stent was shorter, probably because of the smaller inner diameter, and the radioactive seeds exerted no beneficial effect in inhibiting microorganisms.

Screening and Histopathological Characterization of Korean Carrot Lines for Resistance to the Root-Knot Nematode Meloidogyne incognita.

In total, 170 carrot lines developed in Korea were screened for resistance to Meloidogyne incognita race 1 to select parental genetic resources useful for the development of nematode-resistant carrot cultivars. Using the gall index (GI), gall formation was examined on carrot roots inoculated with approximately 1,000 second-stage juveniles of the nematode 7 weeks after inoculation. Sixty-one carrot lines were resistant (GI ≤ 1.0), while the other 109 were susceptible (GI > 1.0) with coefficient of variance (CV) of GI for total carrot lines 0.68, indicating low-variation of GI within the lines examined. The histopathological responses of two carrot plants from resistant and susceptible lines were examined after nematode infection. In susceptible carrots, giant cells formed with no discernible necrosis around the infecting nematodes. In the resistant carrot line, however, no giant cells formed, although modified cells were observed with extensive formation of necrotic layers through their middle lamella and around the infecting nematodes. This suggested that these structural modifications were related to hypersensitive responses governed by the expression of true resistance genes. Therefore, the Korean carrot lines resistant to the nematode infection are potential genetic resources for the development of quality carrot cultivars resistant to M. incognita race 1.

Effect of residual stress on peak cap stress in arteries.

Vulnerable plaques are a subset of atherosclerotic plaques that are prone to rupture when high stresses occur in the cap. The roles of residual stress, plaque morphology, and cap stiffness on the cap stress are not completely understood. Here, arteries are modeled within the framework of nonlinear elasticity as incompressible cylindrical structures that are residually stressed through differential growth. These structures are assumed to have a nonlinear, anisotropic, hyperelastic response to stresses in the media and adventitia layers and an isotropic response in the intima and necrotic layers. The effect of differential growth on the peak stress is explored in a simple, concentric geometry and it is shown that axial differential growth decreases the peak stress in the inner layer. Furthermore, morphological risk factors are explored. The peak stress in residually stressed cylinders is not greatly affected by changing the thickness of the intima. The thickness of the necrotic layer is shown to be the most important morphological feature that affects the peak stress in a residually stressed vessel.

Induction of reparative dentin formation on exposed dental pulp by dentin phosphophoryn/collagen composite.

The ultimate goal of vital pulp therapy is to regenerate rapidly dentin possessing an excellent quality using a biocompatible, bioactive agent. Dentin phosphophoryn (DPP), the most abundant noncollagenous polyanionic protein in dentin, cross-linked to atelocollagen fibrils was applied to direct pulp capping in rats. After 1, 2, and 3 weeks, the teeth applied were examined on the induction of reparative dentin formation and the response of pulp tissue, compared to calcium hydroxide-based agent conventionally used. The reparative dentin formation induced by DPP/collagen composite was more rapid than by calcium hydroxide. In the morphometrical analysis, the formation rate of reparative dentin by DPP/collagen composite was approximately the same as that by calcium hydroxide at 3 weeks. Nevertheless, the compactness of reparative dentin formed by DPP/collagen composite was much superior to what resulted from calcium hydroxide. Also, DPP/collagen composite showed high covering ability of exposed pulp. Moreover, DPP/collagen composite led only to slight pulp inflammation at the beginning whereas calcium hydroxide formed necrotic layer adjacent to the material and induced severe inflammation in pulp tissue at 1 week. The present study demonstrates a potential for DPP/collagen composite as a rapid biocompatible inducer for the formation of reparative dentin of excellent quality in rats.

Gross and histological comparison of hydatid cyst infection in livers of sheep and cows

Three samples of fertile hydatid cysts and 3 other sterile were collected from sheep liver and 3 samples of sterile hydatid cysts from cows liver. Infected liver samples with hydatid cyst were obtained from meat seal shops in AL-Muthanna province in December 2010, then measured dimensions of cysts and calculated the quantity of hydatid fluid and viability of protoscolices by staining with eosin stain then examining under a light microscope of 40x. petechial hemorrhage and paleness around cysts were important gross lesions in sheep in addition to calcification in cow hydatid cysts, on another hand the histological lesions were thickened fibrous layer and severe necrosis with infiltrations of eosinophils and extensive calcification in cows liver. There was a significant effect of the amount of hydatid cyst, thickness of the fibrous layer and thickness of the necrotic layer.

Lesion dimensions following ablative fractional laser treatment in non-melanoma skin cancer and premalignant lesions

Non-melanoma skin cancer (NMSC), which comprises basal cell carcinomas (BCC) and squamous cell carcinomas (SCC), is the most common human cancer with an estimated incidence of more than one million new cancers per year alone in the USA [1–5]. SCC may be preceded by precursor lesions such as actinic keratoses (AK) and SCC in situ (Bowen’s disease), whereas BCCs are regarded as invasive tumours from the beginning. The incidence of the premalignant lesions is even higher with a reported prevalence of AK at 15.4% in men over 40 years of age in a UK population [5]. Ablative fractional laser (AFXL) resurfacing has over the past 5 years gained increasing popularity due to its clinical efficacy and short healing times compared to conventional ablative laser treatment [6]. AFXL creates microscopic vertical channels in the skin, surrounded by undamaged viable tissue. These vertical laser channels, socalled microscopic ablation zones (MAZ), are surrounded by a microscopic treatment zone, which consists of a thin necrotic layer and a coagulated tissue zone [7]. Fractional ablative lasers are available as CO2 lasers (10,600 nm), erbium yttrium aluminium garnet lasers (Er:YAG, 2,940 nm) and yttrium scandium gallium garnet lasers (YSGG, 2,790 nm), all of which have a high absorption in the target chromophore, water, and thus target intraand extracellular water-rich components such as keratinocytes and vessels [6]. AFXL resurfacing is currently used in the treatment of photodamaged skin, wrinkles, and acne and burn scars, but so far not for malignant or premalignant lesions, which is the case for conventional ablative lasers [9–13]. The immediate tissue damage after AFXL and the dimensions of the laser-ablated channels can be characterized histologically [9, 15]. MAZ dimensions have previously been investigated in animal models [14] and normal skin [15–17], whereas no data to our knowledge are available on MAZ dimensions from NMSC and dysplastic lesions. As premalignant and malignant lesions differ histologically from normal skin in several aspects including content of cell types, tumour load, vascularization, and collagen and water content, AFXL may induce different ablation patterns in dysplastic skin and normal skin. Recently, AFXL-assisted drug delivery has been introduced as a novel treatment concept to intensify the biodistribution of topical pharmaceuticals [18]. In vivo studies in pigs have shown that AFXL enhances deep tissue penetration and photoactivation of topically applied methyl aminolevulinate, which is a photosensitizer used for photodynamic therapy (PDT) [18, 19]. PDT is based on photosensitizer uptake by tumour tissue followed by illumination with visible light, which in the presence of oxygen forms cytotoxic oxygen species, causing apoptosis and necrosis of tumour tissue. PDT is approved for the treatment of BCC and premalignant lesions with high cure rates of 90% being reported for superficial malignant lesions, but less effective cure rates of 70–80% for thicker lesions [20]. The inferior treatment outcome in thick lesions K. Togsverd-Bo (*) : C. S. Haak :M. Haedersdal Department of Dermatology, Bispebjerg Hospital, University of Copenhagen, Bispebjerg Bakke 23, 2400 Copenhagen, NV, Denmark e-mail: ktogsverdbo@dadlnet.dk

Histopathology of Experimental Myiasis in Mice as a Result of Infestation and Experimental Implantation of Dermatobia hominis Larvae

A laboratory model of myiasis as a result of Dermatobia hominis (L.) larvae was developed using mice as hosts. Mice in three groups were each infested with one newly hatched larva and skin biopsies processed for histopathology at 4, 12, and 20 d postinfestation (dpi). Mice in three other groups were each subjected to implantation of one larva collected from an infested (donor) mouse at 4, 12, and 20 dpi. Skin lesions of these receptor mice were then assessed at 10, 14, and 6 d postimplantation (dpimp), respectively. The inflammatory process in infested mice at 4 dpi was discrete, consisting of a thin necrotic layer around the larva, edema, many neutrophils, few eosinophils, mast cells, and proliferation of fibroblasts. At 12 dpi, there was a thicker necrotic layer, edema, many neutrophils and eosinophils, few mast cells, neoformation of capillaries, proliferation of the endothelium and fibroblasts, and early stages of fibrosis. These histopathological characteristics together with fibrosis were observed over a large area of the lesion at 20 dpi. Mice submitted to larval implantations demonstrated similar skin histopathology to that seen in the infested rodents, 10 dpimp corresponding to 12 dpi and 6 or 14 dpimp to 20 dpi. In all mice, the progressive acute inflammatory process followed a sequence linked to factors such as size of larvae and presence of secretory-excretory products. Both infested mice and those implanted experimentally with D. hominis larvae were shown to be suitable models for the study of the parasite-host relationship in this important zoonotic myiasis.

A Layered Model for the Growth of Solid Tumors in Vascular Phase

We divide a solid tumor in vascular phase into a vascular layer, an oxygen-diffusion layer, an oxygen-resistant layer and a necrotic layer outside-to-inside, and have constructed a layered model for tumor growth. The actual growth curves of the tumors have been obtained by using different invasion speeds of the vascular layer. The simulative calculation results indicate that the angiogenesis rate is positively correlated with malignant degree of the tumors, if the tumors enter vascular phase, survival time of tumor patients is about 1-4 years, but the tumor patients without angiogenesis can live with tumor.

Rapid and Selective Enzymatic Debridement of Porcine Comb Burns With Bromelain-Derived Debrase®: Acute-Phase Preservation of Noninjured Tissue and Zone of Stasis

Deep burns are associated with the formation of an eschar, which delays healing and increases the risk of infection. Surgical debridement of the eschar is, at present, the fastest means to achieve an eschar-free bed, but the process can not differentiate between the viable tissue and the eschar and follow the minute irregularities of the interface between the two. We evaluated the efficacy and selectivity of a novel enzymatic bromelain-based debriding agent, Debrase Gel Dressing (Debrase), in a porcine comb burn model. We hypothesized that Debrase would result in rapid debridement of the eschar without adverse effects on the surrounding uninjured skin. This is a prospective, controlled, animal experiment. Five domestic pigs (20-25 kg) were used in this study. Sixteen burns were created on each animal's dorsum using a brass comb with four rectangular prongs preheated in boiling water and applied for 30 seconds, resulting in four rectangular 10 x 20 mm full-thickness burns and separated by three 5 x 20 mm unburned interspaces representing the zone of stasis. The burned keratin layer (blisters) was removed, and the burns were treated with a single, topical, Debrase or control vehicle application for 4 hours. The Debrase/control was then wiped off using a metal forceps handle, and the burns were treated with a topical silver sulfadiazine (SSD). The wounds were observed, and full-thickness biopsies were obtained at 4 and 48 hours for evidence of dermal thickness, vascular thrombosis, and burn depth, both within the comb burns and the unburned interspaces in between them. Chi-square and t tests are used for data analysis. A single 4-hour topical application of Debrase resulted in rapid and complete eschar dissolution of all the burns in which the keratin layer was removed. The remaining dermis was thinner (1.1 +/- 0.7 mm) than in the control burns (2.1 +/- 0.3 mm; difference 0.9 mm [95% confidence interval: 0.3-1.4]) and was viable with no injury to the normal surrounding skin or to the unburned interspaces between the burns, which represents the zone of stasis. In control burns, the entire thickness of the dermis was necrotic. At 48 hours, Debrase-treated areas were found partially desiccated under SSD treatment. The unburned interspaces demonstrated partial-thickness necrosis in two third and full-thickness necrosis in one third of wounds. In contrast, full-thickness necrosis was noted in all control interspaces (P = .05). In a porcine comb burn model, a single, 4-hour topical application of Debrase resulted in rapid removal of the necrotic layer of the dermis with preservation of unburned tissues. At 48 hours, SSD treatment resulted in superficial tissue damage and partial preservation of the unburned interspaces.

Possibility of application of calcium carbonate in pulpotomy of rat molars

We investigated restitution processes in mechanically exposed rat molar pulp during pulpotomy with calcium carbonate (CC). The results of the CC treatment were then compared with Calvital ® -containing calcium hydroxide (CH). Micro-computerized tomography (micro-CT), hematoxylin and eosin (H&E) staining and immunoreactivity for nestin, dentin matrix protein-1 (DMP-1) and osteopontin (OPN) were also analyzed. The increment of dentinlike calcified tissue in the pulp was observed by micro-CT. Both CC and CH groups induced pulpotomy resulted in changes associated with inflammation followed by progressive odontoblasts differentiation, dentin matrix secretion and dentin-like bridge formation. Necrotic layer formation and moderate to severe inflammation occurred during the early stages in the CH group. Necrotic layer formation was not observed in the CC group and only associated with mild to moderate inflammation. Immunoreactivity of nestin was observed earlier in the CC group than the CH group. In the CC group, immunoreactivity of DMP-1 was identified beneath the amputated site after 7 days, before increasing until 28 days, and immunoreactivity of OPN was observed in the dentin-like bridge at 28 days, which was also similar to the CH group. These findings suggested that the primary processes of reparative dentinogenesis after pulpotomy with CC may involve natural pulpal wound-healing mechanisms that are similar to the restitution processes observed during pulpotomy with CH. However, CC may prove to be less irritation and more calcified tissue formation than traditional CH-based materials when used as a pulpotomy agent.

COMPATIBILITY STUDIES OF INTERSPECIFIC IN VITRO MICROGRAFTING OF AGARWOOD (Aquilaria malaccensis LAMK.)

Aquilaria spp produced agarwood as nonwood forest production, and has high economic value. A. malaccensis is susceptible to white rot diseases and termites. On the other hand most of the community plantations are a mixed culture with rubber trees, oil palms and with high risk of contamination causing white root diseases. Besides that, vegetative propagation by cuttings, stumping or air layering are still very difficult with low percentage of growth. The objectives of this research were to analyze the best suitable micrograft type, changes of SDS-PAGE protein band patterns of compatible and incompatible micrografts with several combinations of gaharu planlets in in vitro condition, and histology of union area between rootstocks and scion. The results showed that wedge or V type was the best of the micrografs. MS medium with the addition of 3 mg/L IBA was the best medium for gaharu planlet growth after micrografting. Acclimatization was conducted in husk chacoal and top soil (1:1) medium and grown under plastic house of 70% shading with paranet. Compatible combination (Ac/Am) of micrografting showed that anatomy structure of union area is the same as anatomy structure of non micrograftd planlet. While incompatible (Gv/Am) micrografting produced necrotic layer growth from pith and parenchymateous tissues of the wood in union area  along the middle of radial shoot. Recovery period of union area between stocks and scion is initiated by callus formation from the pith and parenchymatous tissues of the wood. Callus will differentiate into mature cells or tissue and become combined phloem and xylem vessels between rootstocks and scion. SDS-PAGE protein band pattern on compatible combination was the same as plants originated from seedlings. While, incompatible combination produced new protein bands with molecular weight around 21 and 30 kD. Key words : Agarwood, Aquilaria spp, micrografting in vitro, incompatible micrografting, SDS-PAGE protein, incompatible histology.

Abstract 1948: Over-expression of HSP27 Provides Chronic Atheroprotection: Reductions in Foam Cell Content and Inflammatory Responses

Introduction: Heat Shock Protein 27 (HSP27) was recently shown by our laboratory to be a biomarker of atherosclerosis and after 4 weeks of a high fat diet (HFD) apoE −/− mice that over-express HSP27 (or apoE −/− HSP27 o/e mice) show a marked attenuation of aortic lesion size compared to apoE −/− mice - but only in females (ATVB 20005; Circ Res in press). Moreover, we demonstrated that serum HSP27 levels were inversely related to atherosclerotic burden, estrogens mediated the release of HSP27 from cells and extracellular HSP27 bound to Scavenger Receptor-A where it could competitively inhibit acLDL uptake. Purpose/Methods: The goal of this study was to determine if and how HSP27 over-expression persistently attenuates atherogenesis in apoE −/− mice fed a HFD for 12 weeks. Results: Compared to female apoE −/− mice, female apoE −/− HSP27 o/e mice showed persistent reductions in aortic (en face) lesion area and aortic sinus cross-sectional lesion area at 12 weeks (e.g., 38% and 39% respectively vs. 35% and 40% after 4 weeks of HFD). Although our original studies revealed that male apoE −/− HSP27 o/e mice had no protection from atherogenesis after 4 weeks of a HFD, in the current study we noted that 12 weeks of a HFD resulted in a 17% reduction in aortic (en face) lesion area and a 24% attenuation in aortic sinus cross-sectional lesion area with HSP27 over-expression. While lesions observed after 4 weeks of HFD consisted mainly of lipid-laden foam macrophages, after 12 weeks of HFD the histopathology was different and included a distinct foam cell layer (or FCL) luminal to a deeper necrotic layer (e.g., with cholesterol crystals). The FCL area was reduced in size in apoE −/− HSP27 o/e vs. apoE −/− mice (e.g., −63% in females, p=0.03; and −48% in males, p=0.02). In vitro, macrophages from apoE −/− HSP27 o/e mice showed a reduction in TNF-α expression compared to apoE −/− mice (e.g., 40% at baseline, and 55% after LPS provocation). Conclusions: Over-expression of HSP27 provides a persistent atheroprotective effect that benefits both sexes. Chronic lesions are not only smaller in size with HSP27 over-expression but show fewer foam cells and macrophages are less pro-inflammatory. These results highlight a potential clinical role for chronic HSP27 modulation for atheroprotection.

Arabidopsis thaliana as a Model System for Graft Union Development in Homografts and Heterografts

Homografting of Arabidopsis thaliana scions on stocks of A. thaliana and heterografting on other species were used to study the compatibility and the ontogeny of graft union formation. Highly compatible homografting with scions of young leafy inflorescence stems was obtained on stocks of inflorescence stems growing from large 3-month-old A. thaliana plants. Histologic analysis revealed four developmental stages of graft union formation in Arabidopsis homografting: (1) development of a necrotic layer, (2) callus proliferation in the grafted scion, (3) differentiation of new vascular tissues within the scion, and (4) a full vascular graft union formation between the scion and the stock. Vascular connections were formed within the callus bridge between rootstocks and scions 15 days after grafting. Heterografts of Arabidopsis on two members of Brassicaceae, cabbage (Brassica) and radish (Raphanus), showed partial incompatible interaction with a lower level of vascular differentiation. Arabidopsis grafting on tomato (Solanaceae) rootstock showed complete incompatibility and limited noncontinuous differentiation of new vascular tissues that did not cross the scion/stock boundary. Although lacking scion/stock vascular connections, Arabidopsis scions grafted onto tomato rootstock flowered and produced seeds. This may indicate some nonvascular functional connections between the two plants, probably of parenchyma cells, further emphasizing the usefulness of Arabidopsis as a model plant for studying various levels of the complicated scion/stock relationships expressed in grafting biology. Experiments with dye transport in the xylem showed that although in general there was an agreement between the histologic study and dye transport, in Arabidopsis homografts water transport frequency was lower than functional and histologic compatability. We conclude that homografting and heterografting of Arabidopsis inflorescence stems is a convenient and reproducible method for studying the fundamental cellular genetic and molecular aspects of grafting biology.

Early recognition of graft compatibility in Uapaca kirkiana Müell Arg. clones, provenances and species

Examination of callus micro-grafts in Uapaca kirkiana Müell Arg. was carried out with the objective of determining early signs of graft compatibility. Leaves from U. kirkiana, U. nitida and Jatropha curcas trees were used for callus induction. Two pieces of callus were co-cultured on Murashige and Skoog (MS) medium with different supplements. Co-cultured calli were embedded in paraffin wax and dissected. The specimens were stained in safranin and fast green before viewing under a light microscope. Results showed that MS medium with 0.1 mg l−1 thidiazuron (TDZ) and 0.5 mg l−1 naphthaleneacetic acid (NAA) or 1.0 mg l−1 dichlorophenoxyacetic acid (2,4-D) and 0.5 mg l−1 NAA was effective for callus induction. There were no necrotic layers at the unions within U. kirkiana clones and provenances, but a differential growth (irregularity) between U. kirkiana and U. nitida co-cultured calli. Phenol deposits were observed at the union interfaces of U. kirkiana combinations and were high on calli derived from mature trees. Phenol deposits were absent at the union of J. curcas heterografts. Necrotic layers developed at the unions of U. kirkiana and J. curcas micro-grafts and indicating an outright graft incompatibility. Accumulation of phenol deposits at the union interfaces inhibited graft compatibility in many U. kirkiana combinations. Callus fusion technique can be used to identify partners with an outright graft incompatibility, especially for distant related plant species.

Effects of Time of Contact and Concentration of Caustic Agent on Generation of Injuries

Solid caustic soda (CS) ingestion levels continue high in Brazil. The aggressiveness of a caustic agent depends, among other factors, on its concentration and time of contact with mucosa. However, the interdependence of these factors in the production of caustic lesion in the esophageal mucosa is not known, especially regarding CS as the strongest corrosive agent. We analyze the effects of concentration and time of contact on the aggressiveness of CS to the esophagus of live animals. One milliliter of CS at concentrations between 1.83% and 73.33% was applied to rats. The solution was kept in contact with the mucosa for 10 to 120 minutes. Internal and external organ aspects were analyzed and the epithelium, submucosa, muscle layer, and adventitia were analyzed microscopically Epithelial necrosis was observed at all concentrations. Among the necrotic layers, the submucosa was observed starting at the 7.33% concentration, and the muscular layer and adventitia were observed at 14.66% concentration. Damage to the pulmonary parenchyma and trachea occurred at 33.66% after 10 minutes, and perforation of the esophagus was observed only after 120 minutes. After 10 minutes, important corrosive lesions installed in the esophageal layers, expanding in depth and superficial extension. The use of heparin had no effect on the production of lesions. Ten minutes were sufficient to provoke necrosis, and longer contact increased the area of necrosis. Solution concentration levels were more important in damage production: 1.83% was sufficient for epithelial necrosis, 7.33% caused submucosal necrosis, and 14.66% muscle and adventitia necrosis; 33.66% solutions caused lung and trachea damage after 10 minutes and esophageal perforation after 120 minutes.

Use of amnion as a graft material in vestibuloplasty: a preliminary report

Objective The aim of this study was to evaluate the clinical use of amnion as a biodegradable graft material for vestibuloplasty. Study design Seven subjects who had been referred for preprosthetic surgery underwent mandibular vestibuloplasty using Clark's technique and amnion as graft material. Fresh amniotic membrane was placed in the area and an acrylic splint was used with soft liner and 0.4-mm wires to cover the surgical site. The area was reexamined after 1 week, 2 weeks, 4 weeks, 3 months, and 6 months. Results A white necrotic soft tissue layer could be seen with underlying hyperemic tissue and an average reduction of 1 to 3 mm in the depth of the labial vestibule after a week. By the end of the second week, the necrotic layer had disappeared, leaving slightly hyperemic mucosal tissue under. By the third week, the graft area could be noticed but the amnion had completely degenerated and disappeared. After 4 weeks, the subjects could be referred for their prosthodontic treatment. The reduction in the depth of the buccal vestibule ranged from 17% to 40% after 6 months' follow-up. Conclusion Amnion might be used as a potential graft material for vestibuloplasty.

In vitro micrografting and the histology of graft union formation of selected species of prickly pear cactus ( Opuntia spp.)

Horizontal and wedge grafts were utilized with micropropagated prickly pear cactus ( Opuntia spp.) to determine the best method for in vitro micrografts. By frequent sampling of the developmental stages of the micrografts we characterized the histological events during graft union formation. Five Opuntia species ( O. streptacantha Lemaire, O. robusta Wendland, O. cochinera Griffiths, O. leucotricha De Candolle, and O. ficus-indica Linné (Miller)) were used as rootstocks and combined with O. ficus-indica to produce homo- and heterografts. Total growth was recorded after 90 days of ex vitro culture to determine the effect of rootstock genotype on scion development. Our results indicate that the easiest and most successful method for micrografting prickly pear cactus was the horizontal graft. Within 28 days vascular connections occurred within the callus bridge between rootstocks and scions in all genetic combinations. Hence, the Opuntia species stock–scion combinations were compatible. Homografts (grafts between same species) grew significantly faster than the heterografts (grafts between different species) after 90 days of ex vitro transfer. Histological observations revealed five developmental stages of graft union formation: (1) development of a necrotic layer, (2) proliferation of callus bridge at the graft interface, (3) differentiation of new vascular cambium, (4) restoration of the new vascular tissue, and (5) restoration of the continuity of external epidermal tissue at the union zone. We conclude that the in vitro horizontal graft is a successful, easy and reliable method to graft micropropagated prickly pear cactus particularly since no contamination or tissue dehydration occurred and no special structures or adhesives were needed for successful graft union formation. This grafting technique has potential use in the commercial production of other cactus species.

Experimental skin lesions from larvae of the bot fly Dermatobia hominis.

Skin biopsies from larvae of Rattus norvegicus, experimentally infested with Dermatobia hominis (Linnaeus Jr) (Diptera: Cuterebridae), were processed for histopathological studies. Two days after infestation, the first-stage larvae (L1) were located deep in the dermis, surrounded by an inflamed area infiltrated predominantly by neutrophils. On the fourth day a thin necrotic layer could be seen close to the larvae, surrounded by large numbers of neutrophils, lymphocytes, macrophages with a few eosinophils and mast cells. A small warble was formed after the fourth day, increasing in size until the seventh day, when the L1 moulted to the second-stage larva (L2). The inflammatory process continued with increasing numbers of neutrophils, macrophages, lymphocytes, eosinophils and mast cells invading the area, as well as the proliferation of fibroblasts and endothelial cells and the appearance of a few localized haemorrhages. After 18-20 days, the L2 moulted to the third-stage larva (L3), when a few plasma cells could be seen in the inflamed area. At 25-30 days there was a reduction in the necrotic layer, as well as in the number of neutrophils and lymphocytes, although large amounts of eosinophils, plasma cells, and collagen fibres were seen. The L3 usually left the host after 30 days. Two days later, the larval cavity was reduced, mast cells infiltrated the region and collagen fibre production were increased. After 7 days, an intense infiltration of plasma cells and scattered necrotic areas could be seen. A scar formed after 10 days. This study showed the laboratory rat to be a suitable model for studies of D. hominis infestation.

Immunohistochemical study of dental pulp applied with 4-META/MMA-TBB adhesive resin after pulpotomy

The purpose of this study was to investigate nerve regeneration and proliferative activity in amputated pulp tissue after the application of 4-META/MMA-TBB adhesive resin (4-META resin). Calcium hydroxide was used as a control material. At 3 days, fibroblast-like cells were positive for proliferating cell nuclear antigen (PCNA) in both 4-META resin- and calcium hydroxide-treated groups and were located mainly within 0.5 mm from the cut surface. Only a few fragmented neurofilament protein (NFP)-positive nerve fibers were observed in this area. At 7 and 14 days, the number of PCNA-positive cells had gradually decreased and regenerated NFP-positive nerve fibers were observed close to the cut surface of the pulp in both groups. At 21 days in the experimental group, several PCNA-positive cells were still found in the area 0.5 mm from the cut surface, and NFP-positive nerve fibers were detected about 0.15–;0.2 mm from the cut surface. In contrast, a dentin bridge was produced under the necrotic layer at 21 days in the control group. PCNA-positive cells were not found underneath the dentin bridge, but NFP-positive nerve fibers had regenerated close to it. These results suggest that although cell differentiation and nerve regeneration are delayed, wound healing occurred even after the application of 4-META resin to exposed pulp surface the same as calcium hydroxide application. © 2000 John Wiley & Sons, Inc. J Biomed Mater Res, 51, 241–248, 2000

Immunohistochemical study of dental pulp applied with 4-META/MMA-TBB adhesive resin after pulpotomy

The purpose of this study was to investigate nerve regeneration and proliferative activity in amputated pulp tissue after the application of 4-META/MMA-TBB adhesive resin (4-META resin). Calcium hydroxide was used as a control material. At 3 days, fibroblast-like cells were positive for proliferating cell nuclear antigen (PCNA) in both 4-META resin- and calcium hydroxide-treated groups and were located mainly within 0.5 mm from the cut surface. Only a few fragmented neurofilament protein (NFP)-positive nerve fibers were observed in this area. At 7 and 14 days, the number of PCNA-positive cells had gradually decreased and regenerated NFP-positive nerve fibers were observed close to the cut surface of the pulp in both groups. At 21 days in the experimental group, several PCNA-positive cells were still found in the area 0.5 mm from the cut surface, and NFP-positive nerve fibers were detected about 0.15-;0.2 mm from the cut surface. In contrast, a dentin bridge was produced under the necrotic layer at 21 days in the control group. PCNA-positive cells were not found underneath the dentin bridge, but NFP-positive nerve fibers had regenerated close to it. These results suggest that although cell differentiation and nerve regeneration are delayed, wound healing occurred even after the application of 4-META resin to exposed pulp surface the same as calcium hydroxide application.