Regenerated Nerve Defects with a Nerve Conduit Containing Dental Pulp Stem Cells in Pigs: An Immunohistochemical and Electrophysiological Evaluation.

BACKGROUND: Dental pulp stem cells (DPSCs) present an exciting new tool in the field of peripheral nerve regeneration due to their close embryonic origin. In this study, we examined their potential in pigs, using biodegradable collagen conduits filled with DPSCs. To our knowledge, this is the first time DPCSs are tested for peripheral nerve regeneration in such large animal model. MATERIALS AND METHODS: The second lateral incisor was extracted from every animal's lower jaw and stem cells were isolated and cultured. The collagen nerve conduits containing the DPSCs were subsequently transplanted into the transected fifth and sixth intercostal nerves, while the seventh intercostal nerve was used as a control and no stem cells were added on the respective collagen conduit. RESULTS: A histological examination was performed on the 3rd and 6th postoperative months and showed the gradual development of neural tissue and immunohistochemical expression of neuron-specific enolase. An electrophysiological study was performed on the 6th postoperative month and showed similar potentials between the stem cell infusion region (5 ± 0.04 units) and their proximal stumps (5 ± 0.05 units) and slightly smaller potentials in the respective distal stumps (4 ± 0.045 units). CONCLUSION: The nerves where DPSCs were injected exhibited morphological and functional recovery, in contrast to the control nerves where no recovery was detected; thus, there is a first evidence of the therapeutic potential of DPSCs in peripheral nerve regeneration.

Experimental formation of dentin-like structure in the root canal implant model using cryopreserved swine dental pulp progenitor cells.

OBJECTIVES: The purpose of the present study was to present histological and immunohistochemical evidence showing the regenerative capacity of swine dental pulp stem cells (S-DPSCs) seeded on organic or synthetic scaffolds and implanted as hybrid root implants in the jaw bone of minipigs. METHODS: Immature permanent incisor teeth and unerupted premolars at the early root-forming stage were extracted from three 7-month-old minipigs, and mesenchymal stem/progenitor cells were isolated from dental pulp. Cells were cryopreserved in liquid nitrogen. A year later, new permanent incisor and premolar teeth were extracted; pulp tissue was removed; and pieces of root canals of the extracted teeth, containing collagen or Poly(lactic-co-glycolic acid) scaffolds seeded with the autologous cryopreserved DPSCs, were implanted into the fresh post-extraction socket of the mini pig jaw. The resulting constructs were harvested after 6 and 10 weeks and evaluated by histological and immunohistochemical analyses. RESULTS: Six weeks postoperatively, the central canal space of the root implants showed degrading scaffold material. New extracellular matrix had been deposited in a polar predentin-like pattern on the canal dentinal walls by cuboidal nonpolarized cells. Ten weeks postoperatively, newly formed organic matrix had been consistently deposited on the canal walls. The presence of a continuous layer of polarized cells showing typical columnar morphology adjacent to the newly deposited organic matrix was evident. CONCLUSIONS: The interactions of S-DPSCs with the dentin matrix of roots implanted in the jawbone of minipigs constitute a model to study in vivo organization and differentiation potential of DPSCs.

Attenuation of liver ischemia/reperfusion induced apoptosis by epigallocatechin-3-gallate via down-regulation of NF-kappaB and c-Jun expression.

INTRODUCTION: Hepatic ischemia/reperfusion (I/R) activates Kupffer cells and initiates severe oxidative stress with enhanced production of reactive oxygen species (ROS) and tumor necrosis factor-alpha (TNF-alpha). ROS and TNF-alpha mediate the expression of nuclear factors and kinases, activating the signal transduction pathway, and triggering apoptosis. The aim of our study was to evaluate the potential protective effect of (-)-epigallocatechin-3-gallate (EGCG) administration in inhibition of apoptosis by attenuating the expression of NF-kappaB, c-Jun, and caspase-3 in a model of severe hepatic I/R. MATERIALS AND METHODS: Thirty Wistar rats were allocated into three groups. Sham operation, I/R, and I/R-EGCG 50mg/kg. Hepatic ischemia was induced for 60min by Pringle's maneuver. Malondialdehyde (MDA), myeloperoxidase (MPO), light histology, scanning electron microscopy, terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL), and immunocytochemistry for NF-kappaB, c-Jun, caspase-3, analysis on liver specimens and aspartate (AST), and alanine (ALT) transferases analysis in serum, were performed 120min after reperfusion. RESULTS: Apoptosis as indicated by TUNEL and caspase-3 was widely expressed in the I/R group but very limited in the EGCG treated group. Liver was stained positive for NF-kappaB and c-Jun in the I/R group but failed to be stained positive in the EGCG treated group. MDA, MPO, AST, and ALT showed marked increase in the I/R group and significant decrease in EGCG treated group. Significant alterations of liver specimens were observed by light histology and transmission electron microscopy whilst pretreatment with EGCG resulted in parenchymal preservation. CONCLUSIONS: Administration of EGCG is likely to inhibit I/R-induced apoptosis and protect liver by down-regulating NF-kappaB and c-Jun signal transduction pathways.

Effects of mastic gum Pistacia lentiscus var. Chia on innate cellular immune effectors.

BACKGROUND: The essential oil and Chios mastic gum (CMG) are natural antimicrobial agents currently broadly used in medicine owing to their antimicrobial, antioxidant, and hepatoprotective properties. The aim of this study was to investigate the effect of CMG-extracted arabinogalactan proteins (AGPs/CMG) both in vitro and in vivo, under the presence of Helicobacter pylori neutrophil-activating protein (HP-NAP), on the innate cellular immune effectors (neutrophils activations) comparing H. pylori-infected patients and healthy controls. PATIENTS AND METHODS: The in-vivo effect of AGPs/CMG under the presence of HP-NAP in neutrophil activation was investigated in five H. pylori-infected patients and three healthy volunteers who received 1 g daily consumption of CMG for 2 months. All participants did not receive any immunosuppressive medication before or during the trial; patients with infectious diseases that could modify their immunologic status were excluded. In-vitro studies with pull-down experiments to assess the effect of AGPs/CMG under the presence of HP-NAP on the neutrophil activation were also carried out. Neutrophil activation was estimated by nicotinamide adenine dinucleotide phosphate-oxidase assays and optical microscopy methods by measurement of cytochrome C reduction. RESULTS: Neutrophil activation was reduced when incubated in vitro with HP-NAP (P=0.0027) and AGP plus HP-NAP (P=0.0004) in H. pylori-positive patients who consumed AGP for 2 months. Similar results were also obtained when neutrophils were incubated with AGP plus HP-NAP (P=0.0038) in controls. Pull-down experiments showed a specific binding of AGPs to two membrane proteins of neutrophils, possibly suggesting inhibition of neutrophil activation. CONCLUSION: AGPs/CMG inhibit neutrophil activation in the presence of HP-NAP, playing a crucial role in H. pylori-associated pathologies in gastric mucosa.

Inhibition of intestinal ischemia/repurfusion induced apoptosis and necrosis via down-regulation of the NF-kB, c-Jun and caspace-3 expression by epigallocatechin-3-gallate administration.

Intestinal ischemia/reperfusion (I/R) produces reactive oxygen species (ROS) activating signal transduction and apoptosis. The aim of this study was to evaluate the effect of (-)-epigallocatechin-3-gallate (EGCG) administration in inhibition of apoptosis by attenuating the expression of NF-kB, c-Jun and caspace-3 in intestinal I/R. Thirty male wistar rats were used. Group A sham operation, B I/R, C I/R-EGCG 50 mg/kg ip. Intestinal ischemia was induced for 60 min by clamping the superior mesenteric artery. Malondialdehyde (MDA), myeloperoxidase (MPO), light histology, Fragment End Labelling of DNA (TUNEL), immunocytochemistry for NF-kB, c-Jun and caspace-3 analysis in intestinal specimens were performed 120 min after reperfusion. Apoptosis as indicated by TUNEL and Caspace-3, NF-kB and c-Jun was widely expressed in I/R group but only slightly expressed in EGCG treated groups. MDA and MPO showed a marked increase in the I/R group and a significant decrease in the EGCG treated group. Light histology showed preservation of architecture in the EGCG treated group. In conclusion, EGCG pre-treatment is likely to inhibit intestinal I/R-induced apoptosis by down-regulating the expression of NF-kB, c-Jun and caspase-3.

Attenuation of intestinal ischemia/reperfusion induced liver and lung injury by intraperitoneal administration of (-)-epigallocatechin-3-gallate.

The aim of this study was to evaluate the effect of ( - )-epigallocatechin-3-gallate (EGCG), a natural antioxidant, on liver and lungs after warm intestinal ischemia/reperfusion (I/R). Thirty male Wistar rats were equally divided into a sham-operation group, an intestinal I/R group and an intestinal I/R group pretreated with EGCG intraperitoneally. Intestinal ischemia was induced by occlusion of the superior mesenteric artery for 60 min followed by reperfusion for 120 min. Immediately after reperfusion, liver, lung and blood samples were collected and analyzed. Results showed that intestinal I/R increased the levels of aspartate (AST) and alanine (ALT) transaminase in serum to 987 and 752 IU/l, respectively. Malondialdehyde (MDA) increased in liver to 1.524 nmol/g in the group subjected to intestinal I/R compared to 0.995 nmol/g in the sham operation group. MDA was also increased in lungs to 1.581 nmol/g compared to 0.896 nmol/g in the sham operation group. Myeloperoxidase (MPO) increased in liver, after intestinal I/R, to 5.16 U/g compared to 1.59 U/g in the sham operation group. MPO was also increased in lungs to 3.89 U/g compared to 1.65 U/g in the sham operation group. Pretreatment with EGCG decreased serum levels of AST and ALT to 236 and 178 IU/l, respectively. It also decreased mean MDA levels in liver and lungs to 1.061 and 1.008 nmol/g, respectively, and mean MPO levels in liver and lungs to 1.88 and 1.71 U/g, respectively. Light microscopy and transmission electron microscopy examinations showed significant alteration in liver and lungs and protection of liver and lung parenchyma in the animals treated with EGCG.

Opacification of hydrophilic acrylic intraocular lenses: a clinical and ultrastructural analysis of three explanted lenses.

BACKGROUND: The purpose of our study was to report the clinical and ultrastructural results of the late opacification of three Hydroview intraocular lenses (IOLs) explanted from three patients with late postoperative visual acuity drop. PATIENTS AND METHODS: Three different patients without any systemic or ocular associated pathology presented with decreased visual acuity 12 to 24 months after uneventful phacoemulsification and Hydroview IOL implantation. The lenses were explanted and examined by photon and transmission electron microscopy (TEM), as well as through laboratory chemical analysis using bleaching with H 2 O 2, SDS-electrophoresis and silver staining for protein detection. They were cut in small pieces, fixed with osmium tetroxide 1.5 %, dehydrated through ascending grades of ethanol, and embedded in EPON 812. Ultrathin sections were stained with aqueous solutions of uranyl acetate and lead citrate, and they were examined with JEOL 2000 CX TEM. One normal IOL was also prepared in the same way and served as control. CONTROL: It was transparent and no material on the surface of semithin and thin sections was detected. Cases 1 and 2: A total gray-whitish opacification of these lenses occurred. An amorphous loose material was present on semithin sections in alternation with normal areas. Evaluation with TEM revealed a pattern of network distribution within the lenses material. The latter was continuous and lightly granulated at the periphery, yet irregularly arranged in the center, more intensely stained, leaving areas of normal appearance. After bleaching, no changes in opacification were seen macroscopically. SDS-electrophoresis and silver staining did not detect proteins. Case 3: This lens had a small opacification close to the periphery. CONCLUSIONS: Late opacification of hydrophilic IOLs is not an uncommon phenomenon after uneventful cataract surgery. Although calcified deposits on the surfaces of such lenses have been reported, this is the first study that describes ultrastructural changes in the IOL material itself.