A. Cevik Tufan

Immunohistochemical demonstration of nerve endings in iliolumbar ligament.

Study Design. Immunohistochemical study on fresh cadaver specimens. Objective. Assessment of mechanoreceptor and nociceptor levels and distribution in iliolumbar ligament. Summary and Background Data. The function of iliolumbar ligament and its role in low back pain has not been yet fully clarified. Understanding the innervation of this ligament should provide a ground which enables formation of stronger hypotheses. Methods. Bilateral 30 iliolumbar ligaments of 15 fresh cadavers were included in the study. Morphologic properties were recorded and the ligaments were examined by focusing on 3 main parts: ligament, bone insertions, and tendon body. Assessment of mechanoreceptor and nociceptor levels and their distribution in iliolumbar ligament were performed on the basis of immunohistochemistry using the S-100 antibody specific for nerve tissue. Results. Iliac wing insertion was found to be the richest region of the ligament in terms of mechanoreceptors and nociceptors. Pacinian (type II) mechanoreceptor was determined to be the most common (66.67%) receptor followed by Ruffini (type I) (19.67%) mechanoreceptor, whereas free nerve endings (type IV) and Golgi tendon organs (type III) were found to be less common, 10.83% and 2.83%, respectively. Conclusion. Immunohistochemical staining has shown that iliolumbar ligamen had a rich nerve tissue. Those results indicate that ILL plays an important role in proprioceptive coordination of lumbosacral region alongside its known biomechanic support function. Moreover, the presence of type IV nerve endings suggest that the injury of this ligament might contribute to the low back pain.

Serum or plasma cartilage oligomeric matrix protein concentration as a diagnostic marker in pseudoachondroplasia: differential diagnosis of a family

Pseudoachondroplasia (PSACH) is an autosomal-dominant osteochondrodysplasia due to mutations in the gene encoding cartilage oligomeric matrix protein (COMP). Clinical diagnosis of PSACH is based primarily on family history, physical examination, and radiographic evaluation, and is sometimes extremely difficult, particularly in adult patients. Genetic diagnosis based on DNA sequencing, on the other hand, can be expensive, time-consuming, and intensive because COMP mutations may be scattered throughout the gene. However, there is evidence that decreased plasma COMP concentration may serve as a diagnostic marker in PSACH, particularly in adult patients. Here, we report the serum and/or plasma COMP concentration-based differential diagnosis of a family with affected adult members. The mean serum and/or plasma COMP concentrations of the three affected family members alive (0.69±0.15 and/or 0.81±0.08 μg/ml, respectively) were significantly lower than those of an age-compatible control group of 21 adults (1.52±0.37 and/or 1.37±0.36 μg/ml, respectively; P<0.0001). Bidirectional fluorescent DNA sequencing-based genetic diagnosis of these patients revealed a heterozygous mutation for the nucleotide change 1532A>G in exon 14 of the COMP gene, resulting in a substitution of amino acid 511 from aspartic acid to glycine in COMP. Thus, serum and/or plasma COMP concentration may be suggested as an additional diagnostic marker to aid clinical and radiographic findings in suspected cases of PSACH.

Penicillin-induced epilepsy model in rats: Dose-dependant effect on hippocampal volume and neuron number

This study was designed to evaluate the penicillin-induced epilepsy model in terms of dose-response relationship of penicillin used to induce epilepsy seizure on hippocampal neuron number and hippocampal volume in Sprague-Dawley rats. Seizures were induced with 300, 500, 1500 and 2000IU of penicillin-G injected intracortically in rats divided in four experimental groups, respectively. Control group was injected intracortically with saline. Animals were decapitated on day 7 of treatment and brains were removed. The total neuron number of pyramidal cell layer from rat hippocampus was estimated using the optical fractionator method. The volume of same hippocampal areas was estimated using the Cavalieri method. Dose-dependent decrease in hippocampal neuron number was observed in three experimental groups (300, 500 and 1500IU of penicillin-G), and the effects were statistically significant when compared to the control group (P<0.009). Dose-dependent decrease in hippocampal volume, on the other hand, was observed in all three of these groups; however, the difference compared to the control group was only statistically significant in 1500IU of penicillin-G injected group (P<0.009). At the dose of 2000IU penicillin-G, all animals died due to status seizures. These results suggest that the appropriate dose of penicillin has to be selected for a given experimental epilepsy study in order to demonstrate the relevant epileptic seizure and its effects. Intracortical 1500IU penicillin-induced epilepsy model may be a good choice to practice studies that investigate neuroprotective mechanisms of the anti-epileptic drugs.

Implication of C-type natriuretic peptide-3 signaling in glycosaminoglycan synthesis and chondrocyte hypertrophy during TGF-β1 induced chondrogenic differentiation of chicken bone marrow-derived mesenchymal stem cells.

This study investigated the involvement of CNP-3, chick homologue for human C-type natriuretic peptide (CNP), in TGF-β1 induced chondrogenic differentiation of chicken bone marrow-derived mesenchymal stem cells (MSCs). Chondrogenic differentiation of MSCs in pellet cultures was induced by TGF-β1. Chondrogenic differentiation and glycosaminoglycan synthesis were analyzed on the basis of basic histology, collagen type II expression, and Alcian blue staining. Antibodies against CNP and NPR-B were used to block their function during these processes. Results revealed that expression of CNP-3 and NPR-B in MSCs were regulated by TGF-β1 in monolayer cultures at mRNA level. In pellet cultures of MSCs, TGF-β1 successfully induced chondrogenic differentiation and glycosaminoglycan synthesis. Addition of CNP into the TGF-β1 supplemented chondrogenic differentiation medium further induced the glycosaminoglycan synthesis and hypertrophy of differentiated chondrocytes in these pellets. Pellets induced with TGF-β1 and treated with antibodies against CNP and NPR-B, did show collagen type II expression, however, Alcian blue staining showing glycosaminoglycan synthesis was significantly suppressed. In conclusion, CNP-3/NPR-B signaling may strongly be involved in synthesis of glycosaminoglycans of the chondrogenic matrix and hypertrophy of differentiated chondrocytes during TGF-β1 induced chondrogenic differentiation of MSCs.

C-type natriuretic peptide regulation of limb mesenchymal chondrogenesis is accompanied by altered N-cadherin and collagen type X-related functions

AMDM, a form of osteochondrodysplasia, is due to the loss‐of‐function mutations in NPR‐B gene. This study investigated the functional involvement of CNP‐3, chick homolog of human CNP, and its receptor NPR‐B in chondrogenesis utilizing the micromass culture of the chick limb mesenchymal cells. Results revealed CNP‐3 and NPR‐B expression in the chick limb bud making stage‐specific peak levels first at Hamburger‐Hamilton stage 23–24, and second at stage 30–31, corresponding to pre‐chondrogenic mesenchymal condensation and initiation of chondrogenic maturation‐hypertrophy in vivo, respectively. CNP‐3 and NPR‐B expression in vitro increased parallel to collagen type X expression, but not to that of collagen type II. Treatment of cultures with CNP significantly increased N‐cadherin, and collagen type X expression, glycosaminoglycan synthesis and chondrogenesis. Collagen type II expression was not significantly affected. Thus, results implicated CNP‐3/NPR‐B signaling in pre‐chondrogenic mesenchymal condensation, glycosaminoglycan synthesis and late differentiation of chondrocytes in the process of endochondral ossification. J. Cell. Biochem. 105: 227–235, 2008.

A Concise Review on the Use of Mesenchymal Stem Cells in Cell Sheet-Based Tissue Engineering with Special Emphasis on Bone Tissue Regeneration

The integration of stem cell technology and cell sheet engineering improved the potential use of cell sheet products in regenerative medicine. This review will discuss the use of mesenchymal stem cells (MSCs) in cell sheet-based tissue engineering. Besides their adhesiveness to plastic surfaces and their extensive differentiation potential in vitro, MSCs are easily accessible, expandable in vitro with acceptable genomic stability, and few ethical issues. With all these advantages, they are extremely well suited for cell sheet-based tissue engineering. This review will focus on the use of MSC sheets in osteogenic tissue engineering. Potential application techniques with or without scaffolds and/or grafts will be discussed. Finally, the importance of osteogenic induction of these MSC sheets in orthopaedic applications will be demonstrated.

Increased Tunel Positive Cells in CA1, CA2, and CA3 Subfields of Rat Hippocampus Due to Copper and Ethanol Co-Exposure

Copper (Cu) is an essential element for life. However, it is toxic at excessive doses, whereas exposure to ethanol (EtOH) has known to cause morphological changes, degeneration, and neuronal loss in central nervous system. A previous investigation by the authors’ group showed that Cu and EtOH co-treatment cause severe hippocampal neuronal loss in CA1, CA2, and CA3 subfields of rat hippocampus. This study was designed to analyze the possible mechanism(s) of action of this effect. In addition, the possible neurogenesis in response to a potent neurodegenerative treatment in rat hippocampus was analyzed. Results demonstrated that Cu and EtOH induced neuronal loss in rat hippocampus was in correlation with the increased cell death analyzed on the basis of TdT-mediated dUTP nick end labeling (TUNEL) assay. On the other hand, neuronal regenerative activity was detectable in analyzed CA1, CA2, and CA3 subfields of the rat hippocampus analyzed on the basis of 5-bromo-2′-deoxy-uridine (BrdU) labeling assay; however, this activity in treated group was not significantly different from that of control group.