An exploratory study of cell stiffness as a mechanical label-free biomarker across multiple musculoskeletal sarcoma cells.

BACKGROUND: Cancer cells are characterized by changes in cell cytoskeletal architecture and stiffness. Despite advances in understanding the molecular mechanisms of musculoskeletal cancers, the corresponding cellular mechanical properties remain largely unexplored. The aim of this study was to investigate the changes in cellular stiffness and the associated cytoskeleton configuration alterations in various musculoskeletal cancer cells. METHODS: Cell lines from five main sarcoma types of the musculoskeletal system (chondrosarcoma, osteosarcoma, Ewing sarcoma, fibrosarcoma and rhabdomyosarcoma) as well as their healthy cell counterparts (chondrocytes, osteoblasts, mesenchymal stem cells, fibroblasts, skeletal muscle cells) were subjected to cell stiffness measurements via atomic force microscopy (AFM). Biochemical and structural changes of the cytoskeleton (F-actin, ß-tubulin and actin-related protein 2/3) were assessed by means of fluorescence labelling, ELISA and qPCR. RESULTS: While AFM stiffness measurements showed that the majority of cancer cells (osteosarcoma, Ewing sarcoma, fibrosarcoma and rhabdomyosarcoma) were significantly less stiff than their corresponding non-malignant counterparts (p < 0.001), the chondrosarcoma cells were significant stiffer than the chondrocytes (p < 0.001). Microscopically, the distribution of F-actin differed between malignant entities and healthy counterparts: the organisation in well aligned stress fibers was disrupted in cancer cell lines and the proteins was mainly concentrated at the periphery of the cell, whereas ß-tubulin had a predominantly perinuclear localization. While the F-actin content was lower in cancer cells, particularly Ewing sarcoma (p = 0.018) and Fibrosarcoma (p = 0.023), this effect was even more pronounced in the case of ß-tubulin for all cancer-healthy cell duos. Interestingly, chondrosarcoma cells were characterized by a significant upregulation of ß-tubulin gene expression (p = 0.005) and protein amount (p = 0.032). CONCLUSION: Modifications in cellular stiffness, along with structural and compositional cytoskeleton rearrangement, constitute typical features of sarcomas cells, when compared to their healthy counterpart. Notably, whereas a decrease in stiffness is typically a feature of malignant entities, chondrosarcoma cells were stiffer than chondrocytes, with chondrosarcoma cells exhibiting a significantly upregulated ß-tubulin expression. Each Sarcoma entity may have his own cellular-stiffness and cytoskeleton organisation/composition fingerprint, which in turn may be exploited for diagnostic or therapeutic purposes.

Influence of left- and right-side total hip arthroplasty on the ability to perform an emergency stop while driving a car.

OBJECTIVE: To show the possible effect of left- and right-side total hip arthroplasty (THA) on the ability to perform an emergency stop when driving a car. DESIGN: Inception cohort. SETTING: A driving simulator using an actual car cabin, specifically developed for the experiment, was used for testing driving ability. PARTICIPANTS: Patients (N=40; 20 left-side THA/20 right-side THA) were tested preoperatively and in increments of 8 days and 6, 12, and 52 weeks after surgery. INTERVENTIONS: Left- and right-side THA. MAIN OUTCOME MEASURES: Reaction time, movement time, total brake response time (TBRT), and maximum brake force. RESULTS: Eight days postoperatively, measurements on driving performance indicated a slight worsening for all outcome parameters in patients after left-side THA and considerably more worsening in patients after right-side THA. For both patient groups, significant improvements in outcome measures were noted during the 1-year follow-up. Brake force declined significantly in patients with left-side THA (P=.012) and in patients after right-side THA (P<.001). A total of 35% of the patients with right-side THA and 15% with left-side THA could not meet the 600 ms TBRT threshold 6 weeks postoperatively. CONCLUSIONS: Most patients who underwent right-side THA reached their preoperative baseline 6 weeks after surgery. Most of the patients with left-side THA showed no TBRT limitations 8 days postoperatively. Because of the patients' highly individual rehabilitation course and considering the possible consequences of the premature resumption of driving a motor vehicle, individual examination and recommendation are necessary.

Radiographic signs for detection of femoroacetabular impingement and hip dysplasia should be carefully used in patients with osteoarthritis of the hip.

BACKGROUND: During the last years, terms like acetabular retroversion, excessive overcoverage, and abnormal head-neck-junction with the so called "pistol-grip-deformity" has been added to the classical description of hip dysplasia. These anatomical changes could lead to a femoroacetabular impingement (FAI). Both kinds of FAI has been indentified as a main reason for hip pain and progressive degenerative changes leading to early osteoarthritis of the hip. A lot of radiographic criteria on pelvic views have been established to detect classical dysplasia and FAI. The present study was initiated to assess the hypothesis that age and severity of osteoarthritis affect measurements of different radiographic parameters. METHODS: The pelvic radiographs of 1614 patients were measured for head-ratio, CE-angle, roof obliquity, extrusion-index, depth-to-width ratio, CCD-angle, sharp's angle. To evaluate the severity of osteoarthritis of the hip the classification by Kellgren and Lawrence was used. Associations between age and radiographic parameters or severity of osteoarthritis were assessed by Spearman's (ρ) or Kendall's (r) rank correlation coefficient, respectively. RESULTS: 366 (22.7%) patients presented no sign of osteoarthritis, 367 (22.7%) patients presented I° osteoarthritis, 460 (28.5%) patients presented II° osteoarthritis, 307 (19%) III° osteoarthritis and 114 (7.1%) IV° osteoarthritis of the hip. The mean head-ratio of all patients was 1.13 ± 0.26 (0.76 - 2.40), the mean CE-angle 40.05° ± 10.13° (0° - 70°), the mean roof obliquity was 35.27° ± 4.96° (10° - 55°), the mean extrusion-index was 12.99 ± 9.21 (6.20 - 95.2), the mean depth-to-width ratio was 59.30 ± 8.90 (6.30 - 100), the mean CCD-angle was 127.68° ± 7.22° (123° - 162°) and the mean sharp's angle was 9.75° ± 5.40° (1° - 34°) There was a weak association between age and the severity of osteoarthritis of the hips (left: r= 0.291; right: r=0.275; both P<0.001) with higher osteoarthritis levels observable for elderly patients). CONCLUSION: Severity of osteoarthritis has a negative impact on measurements of different radiographic parameters. Therefore - in our opinion - epidemiological studies on prearthrotic deformities should only be performed in healthy adults with no signs of osteoarthritic changes.

Circulating platelet-progenitor cell coaggregate formation is increased in patients with acute coronary syndromes and augments recruitment of CD34+ cells in the ischaemic microcirculation.

AIMS: The aim of the present study was to evaluate the levels of platelet interaction with circulating CD34+ cells in patients with stable angina pectoris (SAP) and acute coronary syndromes (ACS) and to study the functional consequence of coaggregates formation in vitro and in vivo. METHODS AND RESULTS: Platelet binding to circulating progenitor cells was defined by the presence of the platelet-specific marker glycoprotein Ib (CD42b) on the surface of CD34+ cells using flow cytometry. The percentage of CD34+/CD42b+ cell coaggregates was increased in patients with ACS (n = 162), and especially in patients with ST-elevation myocardial infarction (STEMI) (n = 44), compared with patients with SAP (n = 116; P < 0.001). In the ANCOVA analysis, platelet/CD34+ cell coaggregates were independently increased in ACS after adjustment for possible confounders. In a subgroup of our cohort, we also evaluated the levels of CD34+/CD133+/CD42b+ cell coaggregates, which were also significantly increased in ACS, and especially in STEMI (P < 0.05). Platelet/CD34+ cell coaggregates formation correlated with platelet activation (P = 0.001). In a prospective pilot study of patients with AMI (n = 40) using cardiac MRI, patients with increased baseline platelet/CD34+ cell coaggregates presented with a less myocardial infarct size and better left ventricular function at a 3-month follow-up compared with patients with lower coaggregates (P < 0.05 for all). The adhesion of platelet/CD34+ cell coaggregates onto the extracellular matrix and to endothelial monolayer was enhanced compared with CD34+ under high shear rates in vitro (P < 0.05) and within the microcirculation in mice after ischaemia/reperfusion injury as assessed by intravital microscopy (P < 0.05). CONCLUSIONS: These findings imply that circulating platelet/CD34+ cell coaggregate levels are increased in ACS, especially in STEMI, which may be a novel mechanism of domiciliation of CD34+ progenitor cells to the injured microvasculature after acute myocardial infarction.

Desmoid Fibromatosis Fused With a Lipoma in the Upper Arm.

Lipoma, the most common mesenchymal tumor, often appears as a slow-growing mass in the musculoskeletal system (MSK). While generally non-invasive, their location can cause symptoms. Desmoid fibromatosis (DF), a rare and locally aggressive neoplasm, poses challenges in MSK system diagnosis and management due to its infiltrative nature. Despite lacking metastatic potential, DF has a high recurrence rate, classifying it as "intermediate, locally aggressive" in the WHO classification. Collaborative efforts among orthopedic surgeons, radiologists, and pathologists are crucial for accurate diagnosis and treatment planning for all tumors of the MSK system. This case report presents the first documented example of a DF within a lipoma, highlighting the challenges of diagnosing and treating musculoskeletal tumors.

5-Aminolevulinic Acid-mediated Photodynamic Therapy on Primary Bone Tumor and Bone Metastases Cell Lines.

Bone metastases are associated with poor prognosis and low quality of life for the affected patients. Photodynamic therapy (PDT) emerges as a noninvasive therapy that can target local metastatic bone lesions. This paper presents an in vitro method to study the PDT effect in adherent cell lines. To this end, we demonstrate a step-by-step approach to subject both primary (giant cell bone tumor) and human bone metastatic cancer cell lines (derived from a primary invasive ductal breast carcinoma and renal carcinoma) to 5-aminolevulinic acid (5-ALA)-mediated PDT. After 24 h post 5-ALA-PDT irradiation (blue light-wavelength 436 nm), the therapeutic effect was assessed in terms of cell migration potential, viability, apoptotic features, and cellular growth arrest (senescence). Post 5-ALA-PDT irradiation, musculoskeletal-derived cell lines respond differently to the same doses and exposure of PDT. Depending on the extent of cellular damage triggered by PDT exposure, two different cell fates-apoptosis and senescence were noted. Variable sensitivity to PDT therapy among different bone cancer cell lines provides useful information for selecting more appropriate PDT settings in clinical settings. This protocol is designed to exemplify the use of PDT in the context of musculoskeletal neoplastic cell lines. It may be adjusted to investigate the therapeutic effect of PDT on various cancer cell lines and various photosensitizers and light sources.

5-Aminolevulinic Acid-Mediated Photodynamic Therapy Potentiates the Effectiveness of Doxorubicin in Ewing Sarcomas.

Ewing sarcomas (ES) are aggressive primary bone tumors that require radical therapy. Promising low toxicity, 5-aminolevulinic acid (5-ALA)-mediated photodynamic therapy (PDT) could enhance the effectiveness of conventional treatment modalities (e.g., doxorubicin (DOX)), improving, thus, the anti-tumorigenic effects. In this study, we investigated the effects of DOX and 5-ALA PDT alone or in combination on three different human ES cell lines. Cell viability, reactive oxygen species (ROS) production, and cellular stiffness were measured 24 h after PDT (blue light-wavelength 436 nm with 5-ALA) with or without DOX. ES cell lines have a different sensitivity to the same doses and exposure of 5-ALA PDT. DOX in combination with 5-ALA PDT was found to be effective in impairing the viability of all ES cells while also increasing cytotoxic activity by high ROS production. The stiffness of the ES cells increased significantly (p < 0.05) post treatment. Overall, our results showed that across multiple ES cell lines, 5-ALA PDT can successfully and safely be combined with DOX to potentiate the therapeutic effect. The 5-ALA PDT has the potential to be a highly effective treatment when used alone or in conjunction with other treatments. More research is needed to assess the effectiveness of 5-ALA PDT in in vivo settings.

Postoperative complications and oncologic outcomes after multimodal therapy of localized high risk soft tissue sarcoma.

BACKGROUND: Standard therapy for localized high-risk soft tissue sarcoma includes surgical resection and neoadjuvant or adjuvant radiation therapy (± chemotherapy and locoregional hyperthermia). No difference in oncologic outcomes for patients treated with neoadjuvant and adjuvant radiation therapy was reported, whereas side effect profiles differ. The aim of this analysis was to analyse oncologic outcomes and postoperative complications in patients treated with multimodal treatment. METHODS: Oncologic outcomes and major wound complications (MWC, subclassified as wound healing disorder, infection, abscess, fistula, seroma and hematoma) were evaluated in 74 patients with localized high-risk soft tissue sarcoma of extremities and trunk undergoing multimodal treatment, and also separately for the subgroup of lower extremity tumors. Clinical factors and treatment modalities (especially neoadjuvant vs. adjuvant radiotherapy) were evaluated regarding their prognostic value and impact on postoperative wound complications. RESULTS: Oncologic outcomes were dependent on number of high risk features (tumor size, depth to superficial fascia and grading), but not on therapy sequencing (however with higher risk patients in the neoadjuvant group). Different risk factors influenced different subclasses of wound healing complications. Slightly higher MWC-rates were observed in patients treated with neoadjuvant therapy, compared to adjuvant radiotherapy, although only with a trend to statistical significance (31.8% vs. 13.3%, p = 0.059). However, except for wound infections, no significant difference for other subclasses of postoperative complications was observed between neoadjuvant and adjuvant therapy. Diabetes was confirmed as a major risk factor for immune-related wound complications. CONCLUSION: Rates of major wound complications in this cohort are comparable to published data, higher rates of wound infections were observed after neoadjuvant radiotherapy. Tumor localization, patient age and diabetes seem to be major risk factors. The number of risk factors for high risk soft tissue sarcoma seem to influence DMFS. Neoadjuvant treatment increases the risk only for wound infection treated with oral or intravenous antibiotic therapy and appears to be a safe option at an experienced tertiary center in absence of other risk factors.

Assessment of 5-Aminolevulinic Acid-Mediated Photodynamic Therapy on Bone Metastases: An in Vitro Study.

Bone is a frequent site of metastases, being typically associated with a short-term prognosis in affected patients. Photodynamic therapy (PDT) emerges as a promising alternative treatment for controlling malignant disease that can directly target interstitial metastatic lesions. The aim of this study was to assess the effect induced by PDT treatment on both primary (giant cell bone tumor) and human bone metastatic cancer cell lines (derived from a primary invasive ductal breast carcinoma and renal carcinoma). After 24 h post light delivery (blue light-wavelength 436 nm) with 5-aminolevulinic acid, the effect on cellular migration, viability, apoptosis, and senescence were assessed. Our results showed that bone metastasis derived from breast cancer reacted with an inhibition of cell migration coupled with reduced viability and signs of apoptosis such as nuclei fragmentation following PDT exposure. A limited effect in terms of cellular viability inhibition was observed for the cells of giant cell bone tumors. In contrast, bone metastasis derived from renal carcinoma followed a different fate-cells were characterized by senescent features, without a notable effect on cell migration or viability. Collectively, our study illustrates that PDT could act as a successful therapy concept for local tumor control in some entities of bone metastases.

Clubfoot treatment with Ponseti method-parental distress during plaster casting.

BACKGROUND: Clubfoot is one of the most prevalent musculoskeletal congenital defects. Gold standard treatment of idiopathic clubfoot is the conservative Ponseti method, including the reduction of deformity with weekly serial plaster casting and percutaneous Achilles tenotomy. It is well known that parents of children with severe and chronic illnesses are mentally stressed, but in recent studies regarding clubfoot treatment, parents were only asked about their satisfaction with the treatment. Largely unknown is parental distress before and during plaster casting in clubfoot. Therefore, we want to determinate first, how pronounced the parents' worries are before treatment and if they decrease during the therapy. Second, we hypothesized that parents faced with an extreme deformity (high Pirani score), reveal more distress, than parents whose children have a less pronounced deformity (low Pirani score). Therefore, we wanted to investigate whether the Pirani score correlates with the parents' mental resilience in relation to the therapy of the child as a global distress parameter. METHODS: To answer this question, we developed a questionnaire with the following emphases: Physical capacity, mental resilience, motion score, parents score, and child score with point scores 1 (not affected) to 6 (high affected). Subsequently, we interviewed 20 parents whose children were treated with clubfeet and determined the Pirani score of the infants at the beginning (T0) and at the end (TE) of the treatment with plaster casting. RESULTS: High values were obtained in child score (Mean (M) = 3.11), motion score (M = 2.63), and mental resilience (M = 2.25). During treatment, mental resilience improved (p = 0.015) significantly. Spearman correlation coefficient between Pirani score (T0) and mental resilience (T0) is 0.21, so the initial hypothesis had to be rejected. CONCLUSION: The issues of the children are in the focus of parental worries concerning clubfoot treatment, especially the assumed future motion and the assumed ability to play with other children. Particular emphasis should be placed on educating parents about the excellent long-term results in the function of the treated feet especially as this topic shows the greatest parental distress.

Quality of Life, Physical and Mental Status and Contentment of Patients with Localized Soft Tissue or Bone Sarcoma: A Questionnaire Analysis.

Extremity soft tissue and bone sarcomas represent a rare group of bone and connective tissue cancers. In literature, there is little information about psycho-emotional status and impact on quality of life after the diagnosis and treatment of this kind of tumors. The aim of this survey was to define the profile of the patients at risk and their need for psychooncological care. Our self-created questionnaire consists of 71 items related to the individual emotional, mental and physical situation after the diagnosis of soft tissue and bone sarcoma. Sixty-six patients, surgically treated at our department, were included. Only 37.5% of the patients considered themselves to be completely emotional stable. Psychooncological treatment was accepted mostly by female patients, by patients with higher education level and by married patients. Emotional stability and confidence in future were associated with a strong familiar background, with numerous consultations of psychooncological service and also to gender and physical condition. Current quality of life was strongly correlated to physical condition. Thanks to our questionnaire, we disclosed few risk factors for negative emotional outcome after therapy, such as higher age, social isolation, female gender and poor physical status.

Antibacterial potency of V.A.C. GranuFoam Silver(®) Dressing.

INTRODUCTION: V.A.C.(®) GranuFoam™ therapy is regularly used in the surgical therapy of infected wounds and soft tissue injuries. Silver nanoparticles can destroy bacterial cell walls and inhibit enzymes for cell replication. Silver dressings are therefore successfully used for many indications in wound therapy. In this study, we investigated the antimicrobial potency of ionic silver released from the silver-coated V.A.C.(®) GranuFoam™ during vacuum therapy. Silver dressing was exposed to agar plates populated with bacteria to measure silver release. MATERIALS AND METHODS: A total of 15 agar plates colonised with either Staphylococcus aureus populations or with Staphylococcus epidermidis, were loaded with V.A.C. GranuFoam Silver(®) Dressing polyurethane foam (KCI, San Antonio, Texas). Each of 13 pieces of silver-coated foam was applied to an agar plate. Two plates were loaded with conventional black foam without any coating. After connecting to a vacuum pump, the vacuum therapy of the 15 plates lasted 5 days. The zone of inhibition of bacterial growth around the foam was measured daily. Silver release was also determined as a function of time. RESULTS: At each time point, there was evidence of silver in the agar independent of bacterial colonisation. The S. aureus agar showed a consecutive increase in silver concentration from baseline upon 48 h after exposure to the negative pressure of V.A.C. therapy. An increasing mean silver level after 48, 72 and 96 h was measured under V.A.C. therapy with a peak value after 120 h. In contrast, the results from the S. epidermidis plates did not follow a linear pattern. At the beginning of vacuum therapy, we documented a rise in silver concentration. After 48-96h, the silver levels fluctuated. A maximum zone of inhibition in both bacterial colonised plates (S. aureus and S. epidermidis) was found 39 h after the start of the V.A.C. GranuFoam Silver(®) therapy. CONCLUSION: From our results, we confirmed the antimicrobial effect of the silver ions against S. aureus and S. epidermidis under continuous V.A.C. GranuFoam(®) Silver therapy with a negative pressure of 25 mmHg. Furthermore we could quantify the amounts of silver, which were released from the foam under negative pressure as a function of time.

Increased circulating progenitor cells in Alzheimer's disease patients with moderate to severe dementia: evidence for vascular repair and tissue regeneration?

Cerebrovascular dysfunction is a common finding in patients with Alzheimer's disease (AD) and may contribute to cognitive decline. Abundant evidence suggests that vascular and neuronal repair mechanisms are mediated by circulating progenitor cells in vivo. Whether CD34+ and, specifically, CD34+/CD133+ progenitor cells are involved in the pathophysiology of AD is poorly understood so far. In the present study, peripheral blood concentrations of circulating CD34+/CD133+ and CD34+ progenitor cells were measured in 45 AD patients and in 30 healthy elderly controls by flow cytometry. The severity of dementia was assessed by Mini-Mental Status Examination and Clinical Dementia Rating scale. AD patients were stratified into two groups showing mild (n=17) and moderate to severe (n= 28) dementia. In the present study, AD patients with moderate to severe dementia, but not those with mild dementia, showed significantly increased circulating CD34+/CD133+ and CD34+ progenitor cells compared to healthy elderly controls independent of cardiovascular risk factors and medication. In addition, the number of circulating CD34+/CD133+ progenitor cells in AD patients was significantly inversely correlated with cognitive function, age, and plasma levels of SDF-1, the most potent chemokine for progenitor cells. Our findings suggest a stage-dependent upregulation of circulating CD34+/CD133+ and CD34+ progenitor cells in AD patients, which could take part in tissue healing processes of the brain in AD.