Jorma Ryhänen

Biocompatibility of nickel-titanium shape memory metal and its corrosion behavior in human cell cultures

Nickel-titanium alloy (Nitinol) is a metallic biomaterial that has a unique thermal shape memory, superelasticity, and high damping properties. Nitinol is potentially very useful in orthopedic surgery, for example. At present, there are not enough confirmative biocompatibility data available on Nitinol. The aim of our study was to clarify the primary cytotoxicity and corrosion rate of Nitinol in human cell cultures. Comparisons were made with stainless steel (Stst), titanium (Ti), composite material (C), and control cultures with no test discs. Human osteoblasts (OB) and fibroblasts (FB) were incubated for 10 days with test discs of equal size, 6 x 7 mm. The cultures were photographed and the cells counted. Samples from culture media were collected on days 2, 4, 6, and 8, and the analysis of metals in the media was done using flameless atomic absorption spectrophotometry. The proliferation of FB was 108% (Nitinol), 134% (Ti) (p < 0.02), 107% (Stst), and 48% (C)(p < 0.0001) compared to the control cultures. The proliferation of OB was 101% (Nitinol), 100% (Ti), 105% (Stst), and 54% (C) (p < 0.025) compared to the controls. Initially, Nitinol released more nickel (129-87 micrograms/L) into the cell culture media than Stst (7 micrograms/L), but after 2 days the concentrations were about equal (23-5 micrograms/L versus 11-1 micrograms/L). The titanium concentrations from both Nitinol and Ti samples were all < 20 micrograms/L. We conclude that Nitinol has good in vitro biocompatibility with human osteoblasts and fibroblasts. Despite the higher initial nickel dissolution, Nitinol induced no toxic effects, decrease in cell proliferation, or inhibition on the growth of cells in contact with the metal surface.

In vivo biocompatibility evaluation of nickel‐titanium shape memory metal alloy: Muscle and perineural tissue responses and encapsule membrane thickness

Nickel-titanium shape memory alloy (Nitinol) has properties that could be very useful in surgical applications. Thermal shape memory, superelasticity, and high damping properties make such alloys behave differently compared to other implant metals. There has previously been a lack of sufficient evidence on the biocompatibility of Nitinol. The purpose of this study was to evaluate general soft tissue response and biocompatibility to Nitinol in vivo, and to clarify neural and perineural responses, previously unreported. Seventy-five rats were randomized into three groups. Test specimens were implanted into paravertebral muscle and near the sciatic nerve. A comparison was made between Nitinol, stainless steel, and Ti-6Al-4V. The animals were euthanized at 2, 4, 8, 12, and 26 weeks after implantation. General morphologic and histologic observations were made under light microscopy. Semiautomatic computerized image analysis was used to measure the encapsule membrane thickness around the implants. The muscular tissue response to Nitinol was clearly nontoxic, regardless of the time period. The overall inflammatory response to Nitinol was very similar to that of stainless steel and Ti-6Al-4V alloy. There were no necroses, granulomas, or signs of dystrophic soft tissue clacification. The immune cell response to Nitinol remained low. Only a few foreign-body giant cells were present. The detected neural and perineural responses were also clearly nontoxic and nonirritating with Nitinol. No qualitative differences in histology between the different test materials could be seen. At 8 weeks, the encapsule membrane of Nitinol was thicker than that of stainless steel (mean 62 +/- 25 microns vs. 41 +/- 8 microns). At the end of the study, the encapsule thickness was equal to all the materials tested. We concluded that Nitinol had good in vivo biocompatibility after intramuscular and perineural implantation in rats in the 26-week follow-up. Based on the results of the present study, Nitinol appears to have good potential for clinical use.

Effect of nickel–titanium shape memory metal alloy on bone formation☆

The aim of this study was to determine the biocompatibility of NiTi alloy on bone formation in vivo. For this purpose we used ectopic bone formation assay which goes through all the events of bone formation and calcification. Comparisons were made between Nitinol (NiTi), stainless steel (Stst) and titanium-aluminium (6%)-vanadium (4%) alloy (Ti-6Al-4V), which were implanted for 8 weeks under the fascia of the latissimus dorsi muscle in 3-month-old rats. A light-microscopic examination showed no chronic inflammatory or other pathological findings in the induced ossicle or its capsule. New bone replaced part of the decalcified matrix with mineralized new cartilage and bone. The mineral density was measured with peripheral quantitative computed tomography (pQCT). The total bone mineral density (BMD) values were nearly equal between the control and the NiTi samples, the Stst samples and the Ti-6Al-4V samples had lower BMDs. Digital image analysis was used to measure the combined area of new fibrotic tissue and original implanted bone matrix powder around the implants. There were no significant differences between the implanted materials, although Ti-6Al-4V showed the largest matrix powder areas. The same method was used for measurements of proportional cartilage and new bone areas in the ossicles. NiTi showed the largest cartilage area (p < or = 0.05). Between implant groups the new bone area was largest in NiTi. We conclude that NiTi has good biocompatibility, as its effects on ectopic bone formation are similar to those of Stst, and that the ectopic bone formation assay developed here can be used for biocompatibility studies.

Bone healing and mineralization, implant corrosion, and trace metals after nickel–titanium shape memory metal intramedullary fixation

Its shape memory effect, superelasticity, and good wear and damping properties make the NiTi shape memory alloy a material with fascinating potential for orthopedic surgery. It provides a possibility for making self-locking, self-expanding, and self-compressing implants. Problems, however, may arise because of its high nickel content. The purpose of this work was to determine the corrosion of NiTi in vivo and to evaluate the possible deleterious effects of NiTi on osteotomy healing, bone mineralization, and the remodeling response. Femoral osteotomies of 40 rats were fixed with either NiTi or stainless steel (StSt) intramedullary nails. The rats were killed at 2, 4, 8, 12, 26, and 60 weeks. Bone healing was examined with radiographs, peripheral quantitative computed tomography, (pQCT) and histologically. The corrosion of the retrieved implants was analyzed by electron microscopy (FESEM). Trace metals from several organs were determined by graphite furnace atomic absorption spectrometry (GF-AAS) or by inductively coupled plasma-atomic emission spectrometry (ICP-AES). There were more healed bone unions in the NiTi than in the StSt group at early (4 and 8 weeks) time points. Callus size was equal between the groups. The total and cortical bone mineral densities did not differ between the NiTi and StSt groups. Mineral density in both groups was lower in the osteotomy area than in the other areas along the nail. Density in the nail area was lower than in the proximal part of the operated femur or the contralateral femur. Bone contact to NiTi was close. A peri-implant lamellar bone sheet formed in the metaphyseal area after 8 weeks, indicating good tissue tolerance. The FESEM assessment showed surface corrosion changes to be more evident in the StSt implants. There were no statistically significant differences in nickel concentration between the NiTi and StSt groups in any of the organs. NiTi appears to be an appropriate material for further intramedullary use because it has good biocompatibility in bone tissue.

Behaviour of Nitinol in osteoblast-like ROS-17 cell cultures

Nickel titanium shape memory metal alloy Nitinol (NiTi) has been used in dental wares and in gastrointestinal surgery. Nitinol is a promising implant material in orthopedics, but its biocompatibility, especially in long-term implantation is not confirmed yet. We studied Nitinols effect on a cell culture model. Comparisons to stainless steel, pure titanium and pure nickel were performed. The effects of Nitinol on cell death rate, the apoptosis rate and the formation of local contacts were studied on rat osteosarcoma cell line ROS-17 in 48-h cultures. The cell death rate was assessed with combined calcein-ethidium-homodimer labelling. The amount of dead cells 1000 cells were as follows: four in the NiTi, 21 in the Stst, 4.8 in the Ti and 51 in the Ni group. In the NiTi and Ti groups, the number of dead cells was significantly lower (p < or = 0.01) than in Ni group. The rate of apoptosis was detected with TUNEL-assay. The assay results were: 1.93 apoptotic cells 1000 cells in the NiTi, 1.1 in the Stst, 2.98 in the Ti and 0.62 in the Ni group. A comparison of these two results shows that 48% of the dead cells were apoptotic in the NiTi, 56.6 in the Stst, 62% in the Ti and only 1.8% in the Ni group. The focal contacts were stained with a paxillin antibody and counted. There were marked differences in the number of focal contacts per unit area compared to NiTi (774 focal contacts): 335 in Stst (p < or = 0.01), 462 in Ti (p < or = 0.01) and 261 in Ni (p < or = 0.005). Our results show that NiTi is well tolerated by the osteoblastic type ROS-17 cells.

Bone modeling and cell–material interface responses induced by nickel–titanium shape memory alloy after periosteal implantation

The purpose of this study was to evaluate the new bone formation, modeling and cell-material interface responses induced by nickel-titanium shape memory alloy after periosteal implantation. We used a regional acceleratory phenomenon (RAP) model, in which a periosteal contact stimulus provokes an adaptive modelling response. NiTi has thermal shape memory and superelasticity properties uncommon in other implant alloys. So far, there are insufficient data concerning the biocompatibility of NiTi as a bone implant. NiTi was compared to stainless steel (stst) and Ti-6Al-4V. The test implant was placed in contact with the intact femur periosteum, but it was not fixed inside the bone. Histomorphometry with digital image analysis was used to determine the bone formation and resorption parameters. The ultrastructural features of cell-material adhesion were analysed with scanning electron microscopy (FESEM). A typical peri-implant bone wall modelation was seen due to the normal RAP. The maximum new woven bone formation started earlier (2 weeks) in the Ti-6Al-4V group than in the NiTi (P < 0.01) group, but also decreased earlier, and at 8 weeks the NiTi (P < 0.05) and stst (P < 0.005) groups had greater cortical bone width. At 12 and 26 weeks no statistical differences were seen in the histomorphometric values. The histological response of the soft tissues around the NiTi implant was also clearly non-toxic and non-irritating. Cell adhesion and focal contacts were similar between the materials studied by FESEM. We conclude that NiTi had no negative effect on total new bone formation or normal RAP after periosteal implantation during a 26-week follow-up.

Unique microRNA profile in Dupuytren's contracture supports deregulation of β -catenin pathway

Dupuytrens contracture, a proliferative disease of unknown origin, is characterized by an abnormal fibroblast proliferation process. Evidence from numerous microRNA (miRNA) studies shows that miRNAs have a vital function in many biological processes, for instance, in cellular signaling networks, cell growth, tissue differentiation, and cell proliferation. Our aim was to characterize, to our knowledge for the first time, the miRNA-expression profile of Dupuytrens contracture. The miRNAs identified may have a function in the pathogenesis of Dupuytrens contracture by targeting and regulating important pathways. We compared the miRNA-expression profile of 29 Dupuytrens contracture patients with that of control samples (fibroblast cells and palmar fascia). Some of the miRNAs identified in our Dupuytrens contracture samples, including miR-29c, miR-130b, miR-101, miR-30b, and miR-140-3p, were found to regulate important genes related to the β-catenin pathway: WNT5A, ZIC1, and TGFB1. Expression profiles of these genes reanalyzed from published gene-expression data from similar patient material correlated with our miRNA results. Analysis was also performed for groups of patients with recurrent/non-recurrent and patients with hereditary/non-hereditary Dupuytrens contracture, but no significant differences appeared in miRNA-expression profiles of these groups. Identification of unique miRNA expression in Dupuytrens contracture may lead to the development of novel molecular therapy for its treatment.

Femoral Neck Is a Sensitive Indicator of Bone Loss in Immobilized Hind Limb of Mouse

The present study was carried out to evaluate a unilateral hind limb immobilization model in the mouse. The right legs of male mice (age 10–12 weeks) were immobilized for 3 weeks against the abdomen by an elastic bandage. Body weight decreased significantly during the immobilization. Peripheral quantitative computed tomography (pQCT) analysis showed that the cross‐sectional cortical area (CSA), the bone mineral content (BMC), and the bone mineral density (BMD) of the tibial diaphysis were lower in both legs of the immobilized animals than in age‐matched controls, but the difference was mainly due to weight reduction. At the tibial metaphysis, CSA, BMC, and BMD were reduced in both legs of the immobilized animals, even after weight adjustment. At the femoral neck, CSA, BMC, and BMD were significantly lower in both legs of the immobilized animals, and the difference between the hind legs of the immobilized animals was also highly significant. The findings of the pQCT study were in good agreement with the changes in mechanical strength. The tibia was a more sensitive indicator of diaphyseal bone weakening than the femur when measuring the bending breaking force of the diaphysis. The femoral neck showed significantly decreased strength, and the difference between the immobilized leg and the contralateral leg was most clearly seen in lateral loading. We conclude that 3 weeks of hind limb immobilization weakened the tibia and femur significantly compared with their contralateral counterparts. The reduction was more significantly seen in the mechanical bending strength than in the pQCT evaluation, and the femoral neck was the most sensitive indicator of bone weakening.

TGF-β1 secretion of ROS-17/2.8 cultures on NiTi implant material

The biocompatibility of an orthopedic implant depends on the effect of the implant on bone-forming cells, osteoblasts. Changes in osteoblastic proliferation, maturation and differentiation are important events in ossification that enable monitoring the effect of the implant. Transforming growth factor-beta (TGF-beta) is known to suppress osteoblast proliferation and, on the other hand, to induce the maturation and differentiation of osteoblasts. Moreover, osteoblasts produce TGF-beta, which is embedded in the bone matrix and activated by bone-resorbing osteoclasts. TGF-beta inhibits osteoclastic activity. Here, we show for the first time the effect of nickel titanium shape memory metal (NiTi) on osteoblastic cytokine expression. In this study, we measured the levels of TGF-beta with enzyme-linked immunosorbent assay (ELISA) from a ROS-17/2.8 osteosarcoma cell line cultured on different metal alloy discs. ELISA results were proportioned to total DNA content of the samples. We compared NiTi, to stainless steel (Stst), pure titanium (Ti) and pure nickel (Ni). The TGF-beta1/DNA value in the NiTi group (0.0007 +/- 0.0003) was comparable with those seen in the Stst (0.0008 +/- 0.0001) and Ti (0.0007 +/- 0.0001) groups. The concentration in the Ni group was lower (0.0006 +/- 0.0003), though not statistically significantly so. In addition, the effect of surface roughness on TGF-beta1 production was studied. We compared three different grades of roughness in three differently hot-rolled alloys: NiTi. hot-rolled at 950 degrees C. Ti alloy hot-rolled at 850 degrees C (TiI) and the same Ti alloy hot-rolled at 1,050 degrees C (TiII). We found that increasing roughness of the NiTi surface increased the TGF-beta1 concentration. On the other hand, all roughness groups of TiII showed low levels of TGF-beta1. while a rough TiI surface induced similar TGF-beta1, expression as rough NiTi. Further, these same measurements made with interleukine 6 (IL-6) were found to be under the detection limit in these cultures. We conclude that a rough NiTi surface promotes TGF-beta1 expression in ROS-17/2.8 cells.

A novel treatment of grade III acromioclavicular joint dislocations with a C-hook implant

Introduction: This study evaluates the results of the new surgical treatment of complete acromioclavicular (ac) dislocations using coracoclavicular (cc) fixation with a shape memory metal C-hook implant. Materials and methods: Fifteen patients were prospectively analyzed. They all had a Tossy III ac dislocation due to trauma. The ac ligament was reinserted using a surgical bone anchor, and the position of the joint was restored by fixing it with a C-hook. After 3 months the C-hook was removed. Functional status, symptom severity, X-rays and patient satisfaction were analyzed during clinical control visits. The follow-up time was 1 year. Results: At 12 weeks, full shoulder function had been achieved by 93% of the patients. The final control visit showed full recovery of active ROM in all patients. Two patients had mild pain during certain movements. X-rays showed the precise anatomical position of ac joint with no statistically significant differences compared to the healthy side. Patient contentment was excellent in 14 cases and satisfactory in one case. The average sick-leave was 58 days, including the removal operation. Minor osteolysis of the clavicle was noticed in two patients. Conclusion: The new C-hook implant provides accurate anatomical reduction, conserves the articular surfaces and enables fast functional recovery with excellent patient contentment. Technically, the implant is easy to use. Based on this study, the C-hook presents a reliable novel treatment option in surgical ac repair.