Diagnostic Value of Frequently Implemented Provocative Tests in the Assessment of Shoulder Pain-A Glimpse of Current Practice.

Initial evaluation of chronic shoulder disability is a diagnostic challenge due to the anatomic complexity of the shoulder joints. For this purpose, several diagnostic tools utilizing provocative testing exist, but only a few have a reliable basis for their diagnostic value. Therefore, objectively determining the predictive value of these tests in identifying the precise anatomical source for disability-subacromial, intraarticular or other-is essential in order to proceed with further imaging evaluations for final objective diagnoses. Using validated clusters of provocative tests should improve their diagnostic values.

Clinical Outcome of Primary Total Hip Arthroplasty in Patients with Morbid Obesity-Retrospective and Prospective Follow-Up Studies.

Background and objective: There is a general clinical concern on the negative impact of obesity on surgical complications and functional outcomes. We hypothesized that the patients with morbid obesity are exceptionally prone to a significantly increased risk for surgical and short-term complications after primary total hip arthroplasty (THA). We aimed to identify the range of Body Mass Index (BMI) values of patients with a significant risk for lower functional improvement after THA. Materials and methods: In Stage 1 of the study, we conducted a retrospective comparative analysis of the rate of complications and functional outcomes in patients treated by primary THA, with normal weight (BMI 19-25, N = 1205) vs. Class 1 (BMI 26-34, N = 450), Class 2 (BMI 35-39, N = 183), and Class 3 (BMI ≥ 40, N = 47) obese patients. After the statistical similarity rates of complications and 6- and 12-month functional outcomes (by Harris Hip and SF-36 scores) were revealed in Class 1 patients and patients with normal BMI, we conducted the Stage 2 prospective study, by the same comparison protocol, on the cohorts of Class 2 (N = 29) and Class 3 (N = 16) patients compared to the Class 1 patients (N = 37) as controls. Results: Stage 1: There was no difference in surgical complications and function on 6- and 12-month postoperative follow-up (physical and mental) between Class 1 and patients with normal BMI (p > 0.05). Surgical complications were significantly higher in Class 2 (p < 0.05) and Class 3 (p < 0.001) patients. Functional activity on the 12-month follow-up increased significantly in all study groups, but in the Class 3 patients, the functional parameters were significantly lower (0.001). The mental health status on the follow-up was similar in all study groups. Stage 2 study revealed similar to the retrospective study comparison of parameters, except for the significantly lower mental health scores in Class 2 and Class 3 patients (p < 0.05) and functional scores in Class 3 patients (p < 0.05). Conclusion: Although the functional ability increased in all patients, it was significantly lower in Class 3 patients (with morbid obesity). Therefore, the patients with Class 1 and Class 2 obesity should be conceptionally distinguished from Class 3 patients in the decision-making process for a primary THA because of the less favorable functional and mental health improvement in those with morbid obesity (Class 3).

In vitro effect of FGIN-1-27, a ligand to 18 kDa mitochondrial translocator protein, in human osteoblast-like cells.

Ligands of 18 kDa mitochondrial translocator protein (TSPO) differ in their cellular effects. We hypothesize that different TSPO ligands might exert different cellular responses. Therefore, following previous studies that showed different cellular responses to two specific TSPO ligands, PK 11195 and protoporphyrin IX, in human osteoblast-like cells in vitro, we now report the cellular response to another specific TSPO ligand, FGIN-1-27 (10(-5) M) (MW 436 kDa), in order to characterize the effects of each TSPO ligand. We found in primary culture of the human osteoblast-like cells that cell numbers were decreased by an average of 30% (p < 0.001) following exposure to 10(-5) M of FGIN-1-27 in comparison to vehicle controls. Cellular [(18)F]-FDG incorporation and ATP content were suppressed, by an average of 43% (p < 0.001) and 83% (p < 0.001), respectively. Mitochondrial mass and ΔΨm increased by an average of 26% (p < 0.01) and 425% (p < 0.0001) respectively. Lactate dehydrogenase activity was enhanced in culture media by 60% (p < 0.05), indicating overall cell death, while no increase in apoptotic levels was observed. Cellular proliferation, as determined by BrdU assay, was not affected. Synthesis of mRNA of TSPO, VDAC 1, and hexokinase 2 decreased in 0.3, 0.3 and 0.5 fold respectively, with accompanying decreases in protein expression of TSPO and Voltage Dependent Anion Channel 1 by 23% (p < 0.001) and 98% (p < 0.001), respectively, but without changes in hexokinase 2 protein expression. Thus it appears that 10(-5) M FGIN-1-27 reduces cell viability, cell metabolism, and mitochondrial function. Previously we found similar effects of PK 11195 on mitochondrial function and cell metabolism and of protoporphyrin IX on cell death in primary osteoblast-like cells.

Short-Term Outcomes of Total Knee Arthroplasty Using a Conventional, Computer-Assisted, and Robotic Technique: A Pilot Clinical Trial.

Background: Total Knee Arthroplasty (TKA) is a prevalent surgical procedure for treating severe knee arthritis, aiming to alleviate pain and restore function. Recent advancements have introduced computer-assisted (CAS) and robot-assisted (RA-TKA) surgical techniques as alternatives to conventional methods, promising improved accuracy and patient outcomes. However, comprehensive comparative studies evaluating the short-term outcomes and prostheses survivorship among these techniques are limited. We hypothesized that the outcome of RA-TKA and/or CAS- TKA is advantageous in function and prosthesis survivorship compared to manually implanted prostheses. Methods: This prospective controlled study compared the short-term outcomes and prostheses survivorship following TKA using conventional, CAS, and RA-TKA techniques. One hundred seventy-eight patients requiring TKA were randomly assigned to one of the three surgical groups. The primary outcomes were knee function (KSS knee score) and functional recovery (KSS function score), which were assessed before surgery three years postoperatively. Secondary outcomes included prosthesis alignment, knee range of movements, and complication rates. Survivorship analysis was conducted using Kaplan-Meier curves, with revision surgery as the endpoint. Results: While all three groups showed significant improvements in knee function postoperatively (p < 0.001), the CAS and RA-TKA groups demonstrated superior prosthetic alignment and higher survivorship rates than the conventional group (100%, 97%, and 96%, respectively). However, although the RA-TKA group had a maximal 100% survivorship rate, its knee score was significantly lower than following CAS and conventional techniques (mean 91 ± 3SD vs. mean 93 ± 3SD, p = 0.011). Conclusion: The RA-TKA technique offers advantages over conventional and CAS methods regarding alignment accuracy and short-term survivorship of TKA prostheses. Since short-term prosthesis survivorship indicates the foreseen rates of mid- and long-term survivorship, the current data have a promising indication of the improved TKA prosthesis's long-term survivorship by implementing RA-TKA. According to the presented data, although the survival rates were 100%, 97%, and 96% in the three study groups, no clinical difference in the functional outcome was found despite the better mechanical alignment and higher survivorship in the group of patients treated by the RA-TKA.

In vitro catabolic effect of protoporphyrin IX in human osteoblast-like cells: possible role of the 18 kDa mitochondrial translocator protein.

In several pathological conditions, when conversion of Protoporphyrin (PP)IX into heme is impaired, a toxic accumulation of PPIX might occur. PPIX has been found to have affinity to the mitochondrial Translocator Protein 18 kDa. Since it is known that TSPO is abundant in human osteoblast cells, thus we assumed that PPIX can affect cellular functions via interactions with TSPO in these cells. Therefore we aimed to study the metabolic responses of human osteoblast to a high (10⁻5 M) concentration of PPIX in vitro. We found that in primary culture of human osteoblast-like cells cell numbers decreased following exposure to PPIX(10⁻5 M). Cellular [¹8F]-FDG incorporation, mitochondrial mass, ATP content were suppressed, and ΔΨm collapsed. Lactate dehydrogenase activity was enhanced in culture media, indicating overall cell death, while no increase in apoptotic levels was observed. Cellular proliferation was not affected. Protein expression of TSPO, VDAC 1, and hexokinase 2 decreased, although the synthesis of mRNA for hexokinase 2 increased. Thus, PPIX(10⁻5 M) has a cytotoxic effect on human osteoblast-like cell in vitro. Since these cells remain viable following exposure to another TSPO ligand, PK 11195 (10⁻5 M), as observed previously by us, the mode of action of PPIX on osteoblast-like cells is not identical to that of PK 11195. Accordingly pathological accumulation of PPIX may cause necrosis of osteoblasts leading to bone mass loss. We show that this phenomenon is unrelated to iron overload.

Postoperative Pain Control by Local Infiltration Analgesia and Peripheral Nerve Block in Primary Prosthetic Total Knee Arthroplasty.

BACKGROUND AND OBJECTIVE: Postoperative (post-op) pain control has an important impact on post-op rehabilitation. The logistics of its maintenance challenge the effect of peripheral nerve block on post-op pain control, with the risk for post-op complications. We hypothesized that perioperative use of local infiltration analgesia (LIA) is comparable to post-op pain control by peripheral nerve block. MATERIALS AND METHODS: We evaluated three groups of patients treated with primary total knee arthroplasty (TKA) due to symptomatic end-stage osteoarthritis with post-op pain control by LIA (LIA group, n=52), femoral plus sciatic nerve block (FSNB) (FSNB group, n=54), and without local or regional analgesia as controls (Control group, n=53). The primary outcome variable was the post-op pain level intensity as measured by the visual analog scale (VAS). Secondary outcome variables were knee function measured by the Knee Society Score (KSS) and the quadriceps muscle strength recovery profile. RESULTS: Up to 4 hours post-op, pain intensity was significantly lower in FSNB patients (P<0.05). This effect of the peripheral nerve block on the pain level disappeared 6 hours post-op. The LIA and FSNB patients showed a significant decrease in pain intensity on days 2 and 3 post-op (P<0.05) with no mutual differences (P>0.05). This effect disappeared on day 4 post-op (P>0.05). The KSS score showed similar significant improvement of functional abilities (P<0.001) in all three groups. There was no difference in KSS scores among the groups 6 months after surgery (P>0.05). Quadriceps muscle recovery profile was similar in the LIA and Control groups, but significantly poorer in the FSNB group (P<0.001). CONCLUSION: The value of very short-term and improved pain relief of post-op FSNB over LIA of the surgical wound should be carefully weighed against its cost, logistics, and potential complication threat.

In vitro mitochondrial effects of PK 11195, a synthetic translocator protein 18 kDa (TSPO) ligand, in human osteoblast-like cells.

The role of the TSPO in metabolism of human osteoblasts is unknown. We hypothesized that human osteoblast metabolism may be modulated by the TSPO. Therefore we evaluated the presence of TSPO in human osteoblast-like cells and the effect of its synthetic ligand PK 11195 on these cells. The presence of TSPO was determined by [(3)H]PK 11195 binding using Scatchard analysis: Bmax 7682 fmol/mg, Kd 9.24 nM. PK 11195 did not affect significantly cell proliferation, cell death, cellular viability, maturation, [(18)F]-FDG incorporation and hexokinase 2 gene expression or protein levels. PK 11195 exerted a suppressive effect on VDAC1 and caused an increase in TSPO gene expression or protein levels. In parallel there was an increase in mitochondrial mass, mitochondrial ATP content and a reduction in ΔΨm collapse. Thus, it appears that PK11195 (10(-5) M) stimulates mitochondrial activity in human osteoblast-like cells without affecting glycolytic activity and cell death.

Evaluation of the potential complications of surgical wound drainage in primary total hip arthroplasty: a prospective controlled double-blind study.

BACKGROUND: The routine use of postoperative wound drainage following total hip arthroplasty (THA) to avoid the creation of excessive haematomas is controversial because of the potential risk of blood loss and wound infection. METHODS: In a prospective double-blind controlled study, 2 groups of patients with hip joint osteoarthritis were operated with primary THA, 1 with surgical wound negative pressure drainage (Group 1 - 635 patients) and the other without (Group 2 - 527 patients). Postoperative blood haemoglobin and haematocrit levels, the necessity for blood transfusion, values of the potential infection markers (serum C-reactive protein values and erythrocyte sedimentation rate), postoperative pain level (according to the VAS scale) and functional outcome (according to Harris Hip Score [HHS] and SF-36 scores) at 12 months postoperatively were compared. RESULTS: A significantly higher drop in blood haemoglobin and haematocrit values was observed in the Group 1 patients (mean drop of 2.2 gr/dl ± 0.25 vs. 1.6 gr/dl ± 0.35 and mean drop of 16% ± 4.0 vs. 11% ± 3.0, respectively, p < 0.01, paired t test). The need for blood transfusion was significantly higher in the Group 1 patients (4.9% vs. 3.9%, p < 0.05, t test). The severity of pain (VAS scale) on the first day after the operation was significantly lower in the Group 1 patients (p < 0.05, t test), but overall, in both groups it was of low intensity (VAS <3). A similar postoperative wound infection rate was observed in both groups (0.4%). HHS and SF-36 scores were similar in both groups. CONCLUSIONS: The drainage of surgical wounds following primary THA due to hip osteoarthritis has a low added value and might cause an increased requirement for blood transfusion.

Management of Failed Surgery for Anterior Glenohumeral Instability: Synopsis of Clinical Evidence.

Failed surgical treatment of anterior shoulder instability should be treated according to clinical principles similar to primary stabilization by addressing risk factors related to the damaged static glenohumeral stabilizers (labrum, capsule and its components, and bony damage to the humeral head and scapular glenoid). In relatively rare conditions when failed primary surgery involves patients with functionally low demands, conservative treatment by strengthening dynamic muscular stabilizers might be considered; otherwise, surgical revision should be strongly considered aimed at improving quality of life. Although the overall failure rate following primary and revision surgery is expected to be below 4%, it is clear that revision surgery is technically demanding. Therefore, the initial recognition and correction of the exact pathology causing glenohumeral instability is crucial to avoid failure of primary surgery and to facilitate the success of the revision procedure, if necessary.

Safety and efficacy of in vitro generated bone-like material for in vivo bone regeneration - a feasibility study.

Bone-like viable tissue can be generated in vitro by utilizing a combination of inorganic matrix, osteoblasts, osteogenic media and application of adequate mechanical stimulation of the cells. To pursue the proof that the in vitro generated bone-like tissue (BLT) is capable of bridging a critical bone gap in vivo without adverse effects, the in vitro cytotoxicity method (MTT) and murine in vivo model were implemented, by implanting the BLT into calvaria critical bone gap in rats. The endpoints for the evaluation of this concept were histological and radiographic data which should show the effectiveness of this method. We found that there was no cytotoxic effect of the BLT according to the MTT assay and no carcinogenic or other morbid effects of the BLT in vivo (mice experiment, n = 10) The critical gaps in BLT -implanted animals (experimental model with rats) demonstrated full bridging of the calvaria critical bone gap with vascularized woven bone (n = 3) as opposed to animals treated with vehicle material (n = 3), which maintained an open gap without any visible closure, according to gross examination, X-ray imaging and histological analysis. The newly formed bone tissue was characterized by pronounced presence of bone marrow regions and newly formed host blood vessels, a strong indication for functional osseointegration. Therefore, the in vitro generated BLT, which causes bone regeneration in critical gaps, has the translational potential to bridge bone non-union defects, without harmful systemic or cytotoxic effects. These initial feasibility results indicate a high safety profile following in vivo implantation of BLT and its potential clinical ability to be used as autologous bone graft.

Photobiomodulation of human osteoblast-like cells in vitro by low-intensity-pulsed LED light.

Visible light irradiation is an emerging area in regenerative medicine research. We hypothesized that low-intensity-pulsed LED light irradiance may exert photobiomodulatory effects on cultured osteoblast-like cells. To test this hypothesis, we investigated cell proliferation and markers of cell maturation and metabolic activity following pulsed LED irradiance. Monolayer explant cultures of human osteoblast-like cells were exposed four times in 24-h intervals to 2 min of pulsed white LED irradiance of 2.4-2.5 mW·cm-2 and its different spectra of 0.2-0.5 mW·cm-2 (frequency range of 10-40 Hz). Cell proliferation was estimated from microscopic cell counting and cell death by lactate dehydrogenase activity in culture media (measured by a colorimetric method). The early markers of osteoblast maturation and metabolic activity, that is, cellular alkaline phosphatase activity and osteocalcin content, were measured using a colorimetric method and ELISA, respectively. Irradiance of 40 Hz caused the highest increase in cell number (P < 0.01). Osteocalcin content in cells decreased following 40 Hz and 10 Hz irradiance (P < 0.05). The 40 Hz blue range irradiance (diffuse transmittance 420-580 nm, maximal cell irradiance 0.5 mW·cm-2 ) caused a decrease in alkaline phosphatase cellular activity (P < 0.001) and an increase in media osteocalcin content (P < 0.05). The 40 Hz green range (diffuse transmittance 560-650 nm, maximal cell irradiance 0.4 mW·cm-2 ) irradiance caused an increase in the number of cells and in cell death. In summary, pulsed (40 Hz) white light irradiance has photomodulatory effects, with its green range spectrum affecting cell proliferation and cell death, and its blue range spectrum affecting cellular maturation and metabolism. The results indicate a low-intensity threshold of photobiomodulation of osteoblast-like cells in vitro.

In vitro Effects of the Specific Mitochondrial TSPO Ligand Ro5 4864 in Cultured Human Osteoblasts.

The 18 kDa mitochondrial translocator protein (TSPO) ligands (10 µM), e. g., protoporphyrin IX, PK 11195 and FGIN-1-27, have different effects on metabolism and protein expression in human osteoblasts. In this study, we investigated the archetypical TSPO specific ligand Ro5-4864 (10 µM) effect in primary osteoblasts in culture aiming to further understand the TSPO role in these mature metabolically active cells.We found that following exposure to Ro5-4864, cellular [18F]-FDG incorporation and ATP content were reduced by 48% (p<0.001) and 44% (p<0.001), respectively. The mitochondrial membrane potential (ΔΨm) increased by 50% (p<0.01), mRNA synthesis of TSPO and voltage dependent anion channel (VDAC1) decreased both by 70%, the TSPO and VDAC1 protein expression decreased by 80% and 68%, respectively (p<0.001). Ro5 4864 caused a decrease in the proportion of cells in the G1 phase (by 20%, p<0.05), shifting the cell cycle to the S and G2/M phases. Furthermore, 63% decrease in hexokinase 2 protein expression (p<0.001) was found. However, we found no significant effects on hexokinase 2 mRNA expression (by RT-PCR). We also did not see significant changes in mitochondrial mass (MitoTracker Green assay), apoptosis rate (TUNEL assay), overall cell death (LDH assay), cellular proliferation (BrdU assay), cell maturation (cellular alkaline phosphatase assay), and the number of cells in the culture.Therefore, an overall effect of Ro5-4864 exhorts is via pathways related to the mitochondrial activity, which is only partly like PK 11195, but not to the other TSPO ligands.

Improvements in survival of the uncemented Nottingham Total Shoulder prosthesis: a prospective comparative study.

BACKGROUND: The uncemented Nottingham Total Shoulder Replacement prosthesis system (Nottingham TSR) was developed from the previous BioModular shoulder prosthesis taking into consideration the causes of the initial implant's failure. We investigated the impact of changes in the design of Nottingham TSR prosthesis on its survivorship rate. METHODS: Survivorship analyses of three types of uncemented total shoulder arthroplasty prostheses (BioModular, initial Nottingham TSR and current Nottingham TSR systems with 11, 8 and 4 year survivorship data respectively) were compared. All these prostheses were implanted for the treatment of disabling pain in the shoulder due to primary and secondary osteoarthritis or rheumatoid arthritis. Each type of the prosthesis studied was implanted in consecutive group of patients--90 patients with BioModular system, 103 with the initial Nottingham TSR and 34 patients with the current Nottingham TSR system. The comparison of the annual cumulative survivorship values in the compatible time range between the three groups was done according to the paired t test. RESULTS: The 8-year and 11-year survivorship rates for the initially used modified BioModular uncemented prosthesis were relatively low (75.6% and 71.7% respectively) comparing to the reported survivorship of the conventional cemented implants. The 8-year survivorship for the uncemented Nottingham TSR prosthesis was significantly higher (81.8%), but still not in the desired range of above 90%, that is found in other cemented designs. Glenoid component loosening was the main factor of prosthesis failure in both prostheses and mainly occurred in the first 4 postoperative years. The 4-year survivorship of the currently re-designed Nottingham TSR prosthesis, with hydroxylapatite coating of the glenoid baseplate, was significantly higher, 93.1% as compared to 85.1% of the previous Nottingham TSR. CONCLUSION: The initial Nottingham shoulder prosthesis showed significantly higher survivorship than the BioModular uncemented prosthesis, but lower than expected. Subsequently re-designed Nottingham TSR system presented a high short term survivorship rate that encourages its ongoing use.

Functional outcome of surgical treatment of symptomatic nonunion and malunion of midshaft clavicle fractures.

Disability caused by nonunited or malunited fracture of the midshaft clavicle is a rare condition that is expressed by local pain or neurovascular impairment. This condition is usually treated by reduction of the fracture and stable fixation with augmentation by autogenous bone graft. We evaluated the functional outcome in 13 patients who were treated by this method. The mean postsurgical follow-up was 41 months. In all patients, satisfactory osseous union was achieved. Only 46% of the patients returned to their previous professional and recreational activities. There was also evidence that the current Constant scores of the affected shoulders remained significantly lower than those of the normal contralateral side. Ten patients reported various degrees of pain, and only three patients were pain-free. We show that, although solid union after realignment of symptomatic nonunion or malunion of midshaft clavicle fractures is predictable, the patients can remain functionally impaired.

Intraosseous monitoring of drilling in lumbar vertebrae by ultrasound: An experimental feasibility study.

The rationale for this project is to evaluate the efficiency of a novel sonographic method for measurements of interosseous distances. The method utilizes a propagating ultrasonic beam through aqueous milieu which is directed as a jet into a drilled tract. We used a plastic model of human L5 vertebra and ex vivo specimen of L5 porcine vertebra and generated 2 mm in diameter tracts in vertebral pedicles. The tracts were created in the "desired" central direction and in the "wrong" medial and lateral directions. The drilled tracts and the residual, up to opposite cortex, distances were measured sonographically and mechanically and compared statistically. We show that "true" mechanical measurements can be predicted from sonographic measurements with correction of 1-3 mm. The correct central route can be distinguished from the wrong misplaced routes. By using the sonographic measurements, a correct direction of drilling in the pedicle of lumbar L5 vertebra can be efficiently monitored.

Triggering cultured human osteoblast-like cells' maturation by an extremely low magnitude alternating electromagnetic field

Introduction: Alternating and pulsed electromagnetic magnetic fields (AEMF and PEMF) of different amplitudes and frequencies can induce metabolic and proliferative effects in osteoblasts, but there is no clearly directed tendency of these effects. I hypothesize that there are extremely low triggering parameters of alternating electromagnetic field (EMF) intensity, i.e., above the background magnetic field on earth but below the lowest AEMF and PEMF that have been investigated to date (above 0.07 mT and below 0.4 mT) that induce cellular response. Methods: Accordingly, human monolayer explant culture replica were exposed four times in 24-hour intervals to two minutes of 10 kHz AEMF or PEMF (10 Hz pulses at a basic 5 kHz frequency) with a maximal EMF intensity of 0.2 mT for both. Cell proliferation was estimated from microscopic cell counting and cell death by lactate dehydrogenase (LDH) specific activity in culture media (measured using a colorimetric method). The early marker of osteoblast maturation, cellular alkaline phosphatase (AP) specific activity, was measured using a colorimetric method (n=6 for all experiment conditions). Results: No difference was found in cell numbers in the culture samples exposed either to AEMF or PEMF and in the LDH's specific activity in culture media in comparison to the unexposed controls (p>0.05, for both). The cellular AP's specific activity increased significantly only in cell cultures exposed to the 10 kHz AEMF (p=0.011). Conclusions: The triggering for human osteoblast activation for maturation by an extremely low AEMF (10 kHz) is at least 0.2 mT, which is distinct and below the previously found triggering range of a PEMF for proliferation induction. Therefore, application of these EMF parameters in a clinical setup by a separate finetuning of osteoblast proliferation and maturation might have a therapeutic value in enhancing damaged boe regeneration (AU)

Optimal parameters for the enhancement of human osteoblast-like cell proliferation in vitro via shear stress induced by high-frequency mechanical vibration / Parámetros óptimos para la mejora de la proliferación de células similares a osteoblastos humanos in vitro a través del esfuerzo cortante inducido por vibraciones mecánicas de alta frecuencia

Introduction: Biomechanical stimulation of cultured human osteoblast-like cells, which is based on controlled mechanical vibration, has been previously indicated, but the exact mechanical parameters that are effective for cells' proliferation enhancement are still elusive due to the lack of direct data recordings from the stimulated cells in culture. Therefore, we developed a low friction tunable system that enables recording of a narrow range of mechanical parameters, above the infrasonic spectrum, that applied uniformly to human osteoblast-like cells in monolayer culture, aiming to identify a range of mechanical parameters that are effective to enhance osteoblast proliferation in vitro.Methods: Human osteoblast-like cells in explant monolayer culture samples were exposed to mechanical vibration in the 10-70Hz range of frequencies for two minutes, in four 24 hours intervals. Cell numbers in culture, cellular alkaline phosphatase activity (a marker of cell maturation), and lactate dehydrogenase activity in culture media (representing cell death) were measured after the mechanical stimulation protocol application and compared statistically to the control cell cultures kept in static conditions. The cell proliferation was deduced from cell number in culture and cell death measurements.Results: We found that 50-70 Hz of vibration frequency protocol (10-30 μm of maximal displacement amplitude, 0.03g of peak-to-peak acceleration) is optimal for enhancing cells' proliferation(p<0.05), with a parallel decrease of their maturation (p<0.01).Discussion: We detected the optimal mechanical parameters of excitation protocol for induction of osteoblast proliferation in vitro by a mechanical platform, which can be used as a standardized method in the research of mechanotransduction in human osteoblast.(AU)

Introducción: La estimulación biomecánica de células similares a osteoblastos humanos cultivadas, que se basa en vibraciones mecánicas controladas se ha demostrado anteriormente, pero los parámetros mecánicos exactos que son efectivos para la mejora de la proliferación de células aún son difíciles de alcanzar debido a la falta de registros de datos directos de las células estimuladas en cultivo. Por lo tanto, desarrollamos un sistema sintonizable de baja fricción que permite el registro de un rango estrecho de parámetros mecánicos, por encima del espectro infrasónico, que se aplica de manera uniforme a células similares a osteoblastos humanos en cultivo monocapa, con el objetivo de identificar un rango de parámetros mecánicos que son efectivos para mejorar la proliferación de osteoblastos in vitro.Métodos: Se expusieron células similares a osteoblastos humanos en muestras de cultivo de monocapa de explante a vibración mecánica en el rango de frecuencias de 10-70 Hz durante dos minutos, en cuatro intervalos de 24 horas. El número de células en cultivo, la actividad de la fosfatasa alcalina celular (un marcador de maduración celular) y la actividad de la lactato deshidrogenasa en los medios de cultivo (que representa la muerte celular) se midieron después de la aplicación del protocolo de estimulación mecánica y se compararon estadísticamente con los cultivos de células de control mantenidos en condiciones estáticas. La proliferación celular se dedujo del número de células en cultivo y mediciones de muerte celular.Resultados: Encontramos que 50-70 Hz de protocolo de frecuencia de vibración (10-30 μm de amplitud de desplazamiento máxima, 0,03 g de aceleración de pico a pico) es óptimo para mejorar la proliferación de células (p <0,05), con una disminución paralela de su maduración (p <0.01).Discusión: Detectamos los parámetros mecánicos óptimos del protocolo de excitación para la inducción de la proliferación de osteoblastos in vitro mediante una plataforma...(AU)

The role of the cytoskeleton in mechanotransduction in human osteoblast-like cells.

The regulation of osteoblast proliferation is a key factor in maintaining bone mass. The enhancement of this process can be achieved by stimulating the proliferation of these cells. Mechanical stimulation is one of the important enhancing factors, but the exact cellular mechanisms of mechanical stimulation, i.e., mechanotransduction, are unknown. In order to investigate the role of the cytoskeleton components in mechanotransduction for cell proliferation, I compared the total DNA content in cultured replicates of osteoblast-like cells derived from three human donors following their exposure to enhancing mechanical stimulation, with and without added specific microtubular and microfilament polymerization blockers (Colchicin and Cytochalasin D, respectively). The results revealed the essential and unique role of the microtubular component of the cytoskeleton in mechanotransduction for proliferation by showing that Colchicin blocked the expected increase in the DNA content after mechanical stimulation of the cultured replicates without altering the total DNA content in replicates at static conditions. Conversely, a specific blockage of the microfilament polymerization presented uniform cytotoxic effect in both static and biomechanically active environments. Since previous reports indicated the essential role of microfilament polymerization for the osteoblast metabolic activity, the results of this study further support the hypothesis that the mechanotransduction mechanisms for proliferation and metabolic activity are mediated by different intracellular pathways.

Intraosseous monitoring and guiding by ultrasound: a feasibility study.

An efficient method for navigation within bone tissue boundaries is essential for performance of surgical procedures, e.g. without damaging of adjacent vulnerable organs. The application of sonographic measuring methods for this purpose promises to be effective in the ability to distinguish soft trabecular bone from dense cortical bone, owing to an order of magnitude difference in acoustic density between these structures. For this purpose, a specific method was developed that utilizes propagation of a 5 MHz ultrasonic wave through an aqueous milieu. Using this method a 0.2mm resolution in measurements is achieved. This resolution is in an order of magnitude lower than is required for a clinical use. A three-stage experimental approach was adopted: measurements in a cubic "phantom" made of a transparent plastic material, in samples of fresh porcine femora, and in a clinical setting of drilling in the upper and lower jaw during dental implantation surgery in nine patients. Two patterns of the detected ultrasound wave reflections were found: low amplitude reflections from the aqueous surrounding and trabecular bone and highly reflected ultrasound waves from the cortical bone. We show that trabecular and cortical bones are distinguishable by real-time ultrasonic measurement. The distances of the drilled tracts, in the range of 58.0-122.0 mm for the "phantom" experiment, 22.6-35.5 mm for the ex vivo experiment and 10.0-11.5 mm in the clinical experiment, and residual distances to the opposite edge of the tested samples and organs, in the range of 21.0-82.0 mm for the "phantom" experiment, 3.8-11.36 mm for the ex vivo experiment and 2.1-6.9 mm in the clinical experiment, were measured by the presented sonographic method and compared statistically, using linear correlation and Bland Altman plot, to the mechanical and/or radiographic measurements in all three stages of the experiment. A correlation coefficient above 0.95 was considered an indication of high correlation, while a value of 0.75-0.94 was considered intermediate, and a value below 0.75 was considered poor. A very high correlation (p < 0.001) and agreement between the sonographic and the "gold standard" measurements techniques, either mechanical or radiographic depending on the experimental setting, were found. Therefore the presented method of intraosseous sonographic measurements may provide an improved method for the monitoring of intraosseous drilling in respect of the currently used mechanical and/or radiographic clinical methods.