Regeneration of Denervated Skeletal Muscles - Brunelli's CNS-PNS Paradigm.

The restoration of voluntary muscle activity in posttraumatic paraplegia in both animal experiments and other clinical applications requires reproducibility of a technically-demanding microsurgical procedure, limited by physicians' understanding of Brunelli's spinal cord grafting paradigm. The insufficient clinical investigation of the long-term benefits of the CNS-PNS graft application warrants additional inquiry. The objective of this study is to explore the potential benefits of the first replicated, graft-induced neuroregeneration of denervated skeletal muscle regarding long-term clinical outcomes and to investigate the effect of Cerebrolysin on neuromodulation. A randomized study evaluating 30 rats, approved by the National Animal Ethics Advisory Committee was performed. The medication was administered postoperatively. For 14 days, 12 rats received Cerebrolysin (serum), 11 received NaCl 0.9% (shams), and 7 were controls. For microsurgery, the lateral corticospinal tract T10 was grafted to the denervated internal obliquus abdominal muscle. On day 90, intraoperative proof of reinnervation was observed. On day 100, 15 rats were euthanized for fixation, organ removal, and extensive histology-morphology examination, and the Wei-Lachin statistical procedure was employed. After an open revision of 16 rats, 8 were CMAP positive. After intravenous Vecuronium application, two (Cerebrolysin, NaCl) out of two rats showed an incomplete compound muscle action potential (CMAP) loss due to glutamatergic and cholinergic co-transmission, while two others showed a complete loss of amplitude. Cerebrolysin medication initiated larger restored muscle fiber diameters and less scarring. FB+ neurons were not observed in the brain but were observed in the Rexed laminae. Brunelli's concept was successfully replicated, demonstrating the first graft induced existence of cholinergic and glutamatergic neurotransmission in denervated grafted muscles. Statistics of the histometric count of muscle fibers revealed larger fiber diameters after Cerebrolysin. Brunelli's CNS-PNS experimental concept is suitable to analyze graft-neuroplasticity focused on the voluntary restoration of denervated skeletal muscles in spinal cord injury. Neuroprotection by Cerebrolysin is demonstrated.

Consensus of clinical neurorestorative progress in patients with complete chronic spinal cord injury.

Currently, there is a lack of effective therapeutic methods to restore neurological function for chronic complete spinal cord injury (SCI) by conventional treatment. Neurorestorative strategies with positive preclinical results have been translated to the clinic, and some patients have gotten benefits and their quality of life has improved. These strategies include cell therapy, neurostimulation or neuromodulation, neuroprosthesis, neurotization or nerve bridging, and neurorehabilitation. The aim of this consensus by 31 experts from 20 countries is to show the objective evidence of clinical neurorestoration for chronic complete SCI by the mentioned neurorestorative strategies. Complete chronic SCI patients are no longer told, "nothing can be done." The clinical translation of more effective preclinical neurorestorative strategies should be encouraged as fast as possible in order to benefit patients with incurable CNS diseases. This manuscript is published as part of the International Association of Neurorestoratology (IANR) special issue of Cell Transplantation.

Altered passive eruption and familial trait: a preliminary investigation.

Altered passive eruption is described as a condition in which the relationship between teeth, alveolar bone, and the soft tissues creates an excessive gingival display and, in turn, in some circumstances, it may reveal a clinical aspect also known as the "gummy smile." The surgical management of such cases is well understood and has been widely described, with mucogingival and osseous resective procedures being predictable surgical means leading to more balanced aesthetics and proper display of the teeth anatomy. The possible familial trait in case of passive eruption and therefore the possibility of recurrence of the same condition in families of siblings or parents of affected patients have been investigated in this study. 20 patients have been selected and treated in both a private practice and university settings and their immediate family trees were evaluated in order to understand the incidence of the condition. 65% of the treated patients had one or more family members with the same condition leading to seeking further investigation on the possible genetic correlation.

Sinus lift tissue engineering using autologous pulp micro-grafts: A case report of bone density evaluation.

BACKGROUND: Although autografts are the standard procedure for bone grafting, the use of bone regeneration by means of dental pulp stem cell is an alternative that opens a new era in this field. Rigenera Protocol is a new technique able to provide the surgeon autologous pulp micro-grafts. MATERIALS AND METHODS: At the Department of Oral Surgery, Don Orione Hospital, Bergamo, Italy, one patient underwent sinus lift elevation with pulp stem micro-grafts gentle poured onto collagen sponge. A CT scan control was performed after 4 months and DICOM data were processed with medical imaging software which gives the possibility to use a virtual probe to extract the bone density. Pearson's Chi-square test was used to investigate difference in bone density (BD) between native and newly formed bone. RESULTS: BD in newly formed bone is about the double of native bone. CONCLUSION: This report demonstrated that micro-grafts derived from dental pulp poured onto collagen sponge are a useful method for bone regeneration in atrophic maxilla.

Anatase-based implants nanocoating on stem cells derived from adipose tissue.

PURPOSE: The aim of this study was to investigate the effect of a new anatase coating with antibacterial properties (Bactercline anatase coating [BAC]) on dental implants in the commitment of stem cells derived from adipose tissue to osteoblasts. MATERIALS AND METHODS: Using real-time reverse transcription polymerase chain reaction, the quantitative expression of specific genes, such as transcriptional factors (runx2 and sp7), bone-related genes (spp1, col1a1, col3a1, alpl, and fosl1), and mesenchymal stem cells marker (eng), was examined. RESULTS: BAC caused induction of bone-related genes such as sp7, fosl1, alpl, and spp1. In contrast, the expression of runx2, col3a1, and col1a1 was decreased in stem cells treated with BAC with respect to untreated cells. CONCLUSION: The obtained results are relevant to better understand the molecular mechanism of bone regeneration and as a model for comparing other materials with similar clinical effects.

Evaluation of gene expression in MG63 human osteoblastlike cells exposed to tantalum powder by microarray technology.

Conventional orthopedic implants are composed from titanium. To improve some characteristics (ie, volumetric porosity, modulus of elasticity, frictional modulus), a new porous tantalum biomaterial has been developed and its biocompatibility reported. By using DNA microarrays containing 20,000 genes, several genes whose expression were significantly up- or down-regulated were identified in an osteoblastlike cell line (MG63) cultured with tantalum powder (TP). The differentially expressed genes cover a broad range of functional activities: signaling transduction; transcription; cell cycle regulation, proliferation, and apoptosis; and cytoskeleton formation. To the authors' knowledge, the data reported represent the first genetic portrait of TP.

Osteoplant acts on stem cells derived from peripheral blood.

OBJECTIVES: The osteoplant is an equine, flexible, heterologous, deantigenic, cortical, and spongy bone tissue, totally reabsorbable, used for implantation of bone tissue, to restore skeletal, even weight-bearing structures. However, how the osteoplant alters osteoblast activity to promote bone formation is poorly understood. MATERIALS AND METHODS: To study how the osteoplant induces osteoblast differentiation in mesenchymal stem cells, the expression levels of bone-related genes, and mesenchymal stem cell markers are analyzed, using real time Reverse Transcription-Polymerase Chain Reaction (RT-PCR). RESULTS: The osteoplant causes induction of osteoblast transcriptional factors such as osterix (RUNX2), and of bone-related genes such as osteopontin (SPP1) and osteocalcin (BGLAP). In contrast the expression of ENG (CD105) is significantly decreased in stem cells treated with osteoplant, with respect to untreated cells, indicating the differentiation effect of this biomaterial on stem cells. CONCLUSION: The obtained results can be relevant to better understand the molecular mechanism of bone regeneration and as a model for comparing other materials with similar clinical effects.

Reconstruction of severely atrophic jaws using homografts and simultaneous implant placement: a retrospective study.

In the past decade, several investigators have reported that implants inserted in autografts in the same operation (ie, simultaneously inserted implants [SIIs]) have achieved excellent results. However, no report regarding SIIs placed in fresh frozen bone (FFB) is available. Thus, the authors planned a retrospective study on a series of SIIs placed in homologue FFB (but not immediately loaded) to evaluate their clinical outcome. In addition, a comparison with implants inserted in FFB in a second stage (ie, delayed inserted implants) was performed. Seventeen patients were grafted with FFB, and 48 implants were inserted in the same operation. Implant diameter and length ranged from 3.25 to 4.0 mm and from 10.0 to 15 mm, respectively. Data were compared with 302 implants inserted in FFB in a second operation during the same period in 64 patients. Analyzing SIIs, it was noted that only 3 implants were lost (ie, survival rate [SVR] = 93.7%), and no differences were detected among the studied variables by using lost implants as a predictor of clinical outcome. On the contrary, by using crestal bone resorption around the implant's neck and specific cutoff values, it was possible to demonstrate that prosthetic restoration (ie, removable overdentures) correlated with a statistically significant lower delta insertion abutment junction (ie, reduced crestal bone loss) and thus with a better clinical outcome. By comparing SIIs with implants inserted in a second stage in FFB, a better outcome for delayed implants was demonstrated. Implants inserted simultaneously with FFB grafts had a high survival and success rate. SIIs inserted in FFB can be considered reliable devices, although a higher marginal bone loss is to be expected when fixed prosthetic restorations are used. Implants inserted in a second surgical stage have a better SVR and success rate than SIIs.

Anorganic bovine bone (Bio-Oss) regulates miRNA of osteoblast-like cells.

Bio-Oss (Geistlich) is composed of an organic bovine bone and has been widely used in several bone regeneration procedures during oral surgery. However, how this biomaterial enhances osteoblast activity to promote bone formation is not completely understood. MicroRNAs (miRNAs) represent a class of small, functional, noncoding RNAs of 19 to 23 nucleotides that regulate the transcription of messenger RNAs (mRNAs) in proteins. In this study, the miRNA microarray technique was used to investigate translation regulation in an osteoblast-like cell line (MG63) exposed to Bio-Oss. Nine up-regulated miRNAs (mir-423, mir-492, mir-191, mir-23a, mir-377, mir-494, mir-214, mir-193b, mir-320) and 4 down-regulated miRNAs (mir-27a, mir-24, mir-188, let-7c) were identified. Because each miRNA regulates 100 mRNAs, only mRNAs related to bone formation were analyzed. The vast majority of detected mRNAs are down-regulated, including some homeobox genes (genes that regulate the morphogenesis of an entire segment of the body), such as noggin and EN1. An indirect positive effect was demonstrated on bone morphogenetic protein-4. To the authors' knowledge, the data reported here are the first on translation regulation in osteoblasts exposed to Bio-Oss. This study may be relevant in better understanding the molecular mechanism of bone regeneration and used as a potential tool for analyzing the combined use of cytokines.

A retrospective study on 287 implants installed in resorbed maxillae grafted with fresh frozen allogenous bone.

BACKGROUND: Several studies have been performed to evaluate the clinical outcome of implants inserted into maxillae grafted with autogenous bone but few reports have focused on maxillae grafted with fresh-frozen allogenous bone (FFAB). PURPOSE: The purpose of this study is to retrospectively evaluate the clinical outcome of implants installed in resorbed maxillae augmented with FFAB. MATERIALS AND METHODS: A total of 69 patients whom had been treated with FFAB grafts to their maxillae and implant placement 4 to 6 months later were retrospectively evaluated. Edentulism was total and partial in 22 and 47 cases, respectively. A total of 287 implants of various systems had been used. A life table analysis was performed. Marginal bone loss was calculated in radiographs. RESULTS: Five of the 287 implants were lost, giving a survival rate (SVR) of 98.3% over a mean follow-up time of 26 months. The marginal bone resorption at the implants was 1.68 mm (SD = 0.44) after 1 year and 1.85 mm (SD = 0.98) after 4 years. The cumulative success rate based on defined criteria was 96% in the first year but decreased to 40% at 4 years because of marginal bone loss. The Kaplan-Meier algorithm demonstrated a better outcome for female patients, removable dentures, and total edentulism. No differences were detected among diameters, lengths, and implant site. CONCLUSION: Implants placed in FFAB showed a high SVR similar to that reported in previous studies on maxillae grafted with autogenous iliac crest bone. Although our data point to more marginal bone loss in partially edentulous patients and for fixed prosthetic restorations, the use of FFAB for reconstruction of the atrophic jaw prior to implant placement can be considered as a reliable alternative to autogenous bone.

Glutamatergic reinnervation and assembly of glutamatergic synapses in adult rat skeletal muscle occurs at cholinergic endplates.

After denervation of adult rat abdominal muscles, the postsynaptic apparatus of neuromuscular junctions (NMJs) retains its original architecture and clustering of acetylcholine receptors (AChRs). When descending fibers of the spinal cord are surgically diverted to this muscle by a nerve grafting procedure, supraspinal glutamatergic neurons can innervate muscle fibers and restore motor function; the newly formed NMJs switch from a cholinergic to a glutamatergic-type synapse. We show here that regenerating nerve endings contact the fibers in an area occupied by cholinergic endplates. These NMJs are morphologically indistinguishable from those in controls, but they differ in the subunit composition of AChRs. Moreover, by immunofluorescence and immunoelectron microscopy, new NMJs express glutamatergic synapse markers. The alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor subunit GluR1 partially colocalizes with AChRs, and vesicular glutamate transporter 2 is localized in the presynaptic compartment. Immunoprecipitation analysis of membranes from reinnervated muscle showed that AMPA receptor subunits GluR1 and GluR2 coimmunoprecipitate with rapsyn, the AChR-anchoring protein at the NMJ. Taken together, these results indicate that cholinergic endplates can be targeted by new glutamatergic projections and that the clustering of AMPA receptors occurs there.

Platform switching and bone platform switching.

Bone platform switching involves an inward bone ring in the coronal part of the implant that is in continuity with the alveolar bone crest. Bone platform switching is obtained by using a dental fixture with a reverse conical neck. A retrospective study was performed to evaluate the effectiveness of conventional vs reverse conical neck implants. In the period between May 2004 and November 2007, 86 patients (55 females and 31 males; median age, 53 years) were operated and 234 implants were inserted: 40 and 194 were conventional vs reverse conical neck implants, respectively. Kaplan-Meier algorithm and Cox regression were used to detect those variables associated with the clinical outcome. No differences in survival and success rates were detected between conventional vs reverse conical neck implants alone or in combination with any of the studied variables. Although bone platform switching leads to several advantages, no statistical difference in alveolar crest resorption is detected in comparison with reverse conical neck implants. We suppose that the proximity of the implant abutment junction to the alveolar crestal bone gives no protection against the microflora contained in the micrograph. Additional studies on larger series and a combination of platform switching and bone platform switching could lead to improved clinical outcomes.

CaPO4 blasted implants inserted into iliac crest homologue frozen grafts.

In the last decade, some investigations have reported that the resorbable blast media surface (also named CaPO4 blasted implants [CaPO4-Bls]) has achieved excellent results. However, no report regarding CaPO4-Bls inserted into fresh frozen bone (FFB) is available. Thus, we planned a retrospective study on a series of CaPO4-Bls inserted into FFB to evaluate their clinical outcome. In the period between December 2003 and December 2006, 16 patients (10 females and 6 males, median age of 55 years) were operated on, and 76 CaPO4-Bls were inserted. The mean implant follow-up was 23 months. Implant diameter and length ranged from 3.25 to 4.5 mm and from 11.5 to 15 mm, respectively. Implants were inserted to replace 7 incisors, 11 cuspids, 31 premolars, and 27 molars. Only 1 out of 76 implants was lost (i.e., survival rate [SVR] = 98.7%), and no differences were detected among the studied variables. When peri-implant crestal bone resorption was used as an indicator of clinical success (i.e., success rate), it was possible to identify some variables that correlated with a better clinical outcome. Specifically, Cox regression showed that removable prosthetic restoration and longer implant length correlated with a statistically significant lower delta implant abutment junction (IAJ; i.e., reduced crestal bone loss) and thus a better clinical outcome. In this study, CaPO4-Bls had high survival and success rates, similar to those reported in previous reports of 2-stage procedures in nongrafted bone. CaPO4-Bls inserted into FFB are reliable devices, although greater marginal bone loss occurs when fixed prosthetic restorations and short implants are used.

Unsuspected plasticity of single neurons after connection of the corticospinal tract with peripheral nerves in spinal cord lesions.

OBJECTIVE: To report an unsuspected adaptive plasticity of single upper motor neurons and of primary motor cortex found after microsurgical connection of the spinal cord with peripheral nerve via grafts in paraplegics and focussed discussion of the reviewed literature. METHODS: The research aimed at making paraplegics walk again, after 20 years of experimental surgery in animals. Amongst other things, animal experiments demonstrated the alteration of the motor endplates receptors from cholinergic to glutamatergic induced by connection with upper motor neurons. The same paradigm was successfully performed in paraplegic humans. The nerve grafts were put into the ventral-lateral spinal tract randomly, without possibility of choosing the axons coming from different areas of the motor cortex. RESULTS: The patient became able to selectively activate the re-innervated muscles she wanted without concurrent activities of other muscles connected with the same cortical areas. CONCLUSION: Authors believe that unlike in nerve or tendon transfers, where the whole cortical area corresponding to the transfer changes its function a phenomenon that we call "brain plasticity by areas", in our paradigm due to the direct connection of upper motor neurons with different peripheral nerves and muscles via nerve grafts motor learning occurs based on adaptive neuronal plasticity so that simultaneous contractions of other muscles are prevented. We propose to call it adaptive functional "plasticity by single neurons". We speculate that this phenomenon is due to the simultaneous activation of neurons spread in different cortical areas for a given specific movement, whilst the other neurons of the same areas connected with peripheral nerves of different muscles are not activated at the same time. Why different neurons of the same area fire at different times according to different voluntary demands remains to be discovered. We are committed to solve this enigma hereafter.

Implants inserted into homografts bearing fixed restorations.

PURPOSE: In the last decade several studies have evaluated the clinical outcome of implants inserted into autografts and rehabilitated with fixed restorations in either one- or two-step surgical protocols. However, no study has investigated implants placed into homografts; thus, a case series analysis was performed to verify the clinical outcome of implants inserted into fresh frozen bone (FFB) and bearing fixed prosthetic restorations. MATERIALS AND METHODS: Fifty-eight patients underwent iliac crest homograft transplants and 238 implants were inserted. Seventy-one double-etched, 19 sandblasted and acid-etched-1 (SLA1), 10 grit-blasted and acid-etched, 73 anodic oxidized, 39 CaPo4 ceramic-blasted, 19 SLA2, and seven additional implants of various types were used. Implant diameter and length ranged from 3 to 5 mm and from 7 to 16 mm, respectively. Implants were inserted to replace 15 incisors, 14 canines, 102 premolars, and 107 molars. A total of 111 restorations were performed. RESULTS: No implants were lost. Cox regression analysis showed that implant type and type of edentulism directly correlated with a lower bone resorption and thus had a better clinical outcome and success rate. CONCLUSION: Implants bearing fixed restorations and inserted into FFB have higher survival and succes rates compared to those placed in nongrafted and grafted jaws reported in previous studies.

A comparison between genetic portraits of normal osteoblasts and osteosarcoma cell lines.

BACKGROUND: Osteosarcoma (OS) is the most frequent malignant bone tumor occurring in young patients in the first two decades of life. Metastases are the cause of 90% of cancer deaths for patients with OS. OS of the jaw is rare and aggressive malignancy constitutes approximately 5-13% of all cases of skeletal OS. Chemotherapy plus surgery are the first choice for treatment. AIMS: Because OS cell lines (OCLs) should share a common pathway with primary OS and new drugs are screened in in vitro systems, new insight about the genetic profiling of OCLs is of paramount importance to a better understanding of the molecular mechanism of this rare tumor and detecting a potential target for specific therapy. MATERIALS AND METHODS: The SAOS2 and TE85 cell lines were analysed using DNA microarrays containing 19,000 genes. Several genes in which expression was significantly differentially expressed in OCLs vs. normal osteoblast (NO) were detected. RESULTS: The differentially expressed genes cover a broad range of functional activities: (a) cell cycle regulation, (b) cell differentiation, (c) apoptosis, and (d) immunity. CONCLUSION: The reported data can be relevant to a better understanding of the biology of OS and as a model for comparing the effect of drugs used in OS treatment.

Mandibles grafted with fresh-frozen bone: an evaluation of implant outcome.

In the last decade several studies have been performed to evaluate the clinical outcome of implants inserted into grafted mandibles with autologous bone, but none is available on mandibles grafted with fresh-frozen bone. Thus, we planned a retrospective study on a series of implants inserted into homologue fresh-frozen bone to evaluate their clinical outcome. Twenty-one patients were operated on, 28 onlay grafts were inserted into the mandible, and 63 implants placed. Patients had total and partial edentulism in 11 and 10 cases, respectively. The mean follow-up was 20 months. No or reduced crestal bone resorption was considered an indicator of success rate to evaluate the effect of several host-, implant-, and occlusal-related factors. The difference between the implant-abutment junction and the bone crestal level was defined as the implant abutment junction (IAJ) and calculated at the time of operation and during follow-up by means of radiographs. Delta IAJ, the difference between theIAJ at the last check-up and the IAJ recorded just after the operation, were stratified according to variables of interests. Kaplan-Meier algorithm and Cox regression were then performed to detect those variables statistically associated with the delta IAJ. Only 2 of 63 implants were lost (i.e., survival rates = 96.8%) and no differences were detected among the studied variables. On the contrary, Cox regression showed that prosthetic restoration (i.e., removable dentures) was the only factor correlated with a statistically significant lower delta IAJ (i.e., reduced crestal bone loss) and thus a better clinical outcome. Implants inserted into mandibles grafted with fresh-frozen bone allografts have high survival rates and success rate, which are comparable with those obtained with autologous iliac crest bone grafts.

Evaluation of clinical prognostic factors in T1 N0 M0 head and neck basal cell carcinoma.

This retrospective study was carried out to assess the clinical outcome of stage I (ie, T1 N0 M0) head and neck basal cell carcinoma (HNBCC) and verify the impact of some clinical parameters on prognosis. The cases of 220 patients with stage I (ie, T1 N0 M0) histologically proven HNBCC were analyzed.The global disease-specific survival rate at 36 months was 100%, irrespective of the type of surgery, and thus, it was demonstrated that radical tumor resection is a viable procedure for stage I HNBCC. However, 11 (5%) of 220 cases had a recurrence. A second procedure was performed in 10 (90.9%) of 11 cases. In 3 of the 10 patients operated on twice (30%), a second recurrence was detected. Thus, a second surgical procedure on a recurring tumor is possible, but the multimodality treatment regimen should be considered for recurring tumors.

Scoring mandibular fractures: a tool for staging diagnosis, planning treatment, and predicting prognosis.

BACKGROUND: Mandibular fractures (MF) are a daily problem in maxillofacial surgery. A staging system for classifying MF is of paramount importance to plan surgery, to define prognosis, and to exchange information among trauma centers. In this article, a classification for MF is proposed. METHODS: The mandible is divided into six sites (symphysis, body, angle, ramus, coronoid, and condyle) and the mandibular nerve divides the alveolar process (i.e., the upper part containing teeth) from the basal bone (placed caudally with respect to the nerve). MF can be summarized using three abbreviations: A = alveolar, B = basal, and C = complete. Consequently, MFs are staged as follows: F in situ = a greenstick fracture; F1 = a single mobile fragment of alveolar or basal bone; F2 = double mobile fractures of the alveolar or basal portion or a single complete separation of the mandibular arch continuity; F3 = a complete double separation of the mandibular arch continuity; F4 = triple or more mandibular arch fragments. To evaluate the suitability of the proposed classification, a retrospective study on a series of 128 patients (and 205 MFs) was performed. Age, gender, stage, clinical diagnosis at admission, type of surgery, and outcome were considered. RESULTS: A good correlation between the proposed classification and the studied variables was detected. CONCLUSIONS: The new classification is a simple and precise method for staging MF. It can summarize MFs and be used in daily practice. It is our understanding, however, that a multicenter study should be performed before the effectiveness of the proposed classification can be clearly stated.