Near-Peer Teaching in Human Anatomy from a Tutors' Perspective: An Eighteen-Year-Old Experience at the University of Bologna.

The University of Bologna School of Medicine in 2003 adopted a near-peer teaching (NPT) program with senior medical students teaching and assisting younger students in human anatomy laboratories. This study aimed to evaluate the effectiveness and outcomes of this program-unique on the Italian academic panorama-from the tutors' perspective. An anonymous online survey was administered to all those who acted as peer tutors in the period from 2003 to 2021; it evaluated tutors' perceptions regarding the influence of the tutoring experience on their skillset gains, academic performance, and professional career. Furthermore, tutors were asked to express their views on the value of cadaver dissection in medical education and professional development. The overall perception of the NPT program was overwhelmingly positive and the main reported benefits were improved long-term knowledge retention and academic performance, improved communication, team-working and time management skills, and enhanced self-confidence and motivation. Most tutors strongly believed that cadaver dissection was an invaluable learning tool in medical education, helped them to develop professionalism and human values, and positively influenced the caring of their future patients. Nearly all the participants highlighted the importance of voluntary body donation for medical education and research. The present results supported the thesis that tutors themselves benefited from the act of teaching peers; this impactful experience equipped them with a wide range of transferable skills that they could draw on as future educators and healthcare professionals.

The whole body donation program at the university of Bologna: A report based on the experience of one of the oldest university in Western world.

Human body dissection is fundamental in medical education, as it allows future physicians to learn about the body's morphology in three dimensions, to recognize anatomical variations and to develop and increase the essential qualities of respect, compassion and empathy for patients. It is equally important in clinical training as it allows surgeons to improve their manual dexterity and practical skills and to test innovative surgical techniques and devices. In Italy prior to 2020, body acquisition and use for study and research purposes were regulated by a generic set of old directives and national decrees which dealt only marginally with these issues. However, in 2013, a whole body donation program was officially set up at the Institute of Human Anatomy of the University of Bologna. Completely free and voluntary informed consent has always been regarded as a core prerequisite and, since its inception, the program exclusively accepted bequeathed bodies. On February 10, 2020, a specific law governing the disposition of post mortem human body and tissues for study, training and scientific research purposes was definitively enacted. The present work traces the University of Bologna's experience leading to the whole body donation program and the brand new dissecting room. It describes the program of Bologna as an example of "good practice" in body donation, aimed at ensuring education and clinical training by means of both traditional gross anatomy and innovative technology. Moreover, it analyzes the results achieved in terms of increased donor enrollment and improved teaching/training quality and the strengths of this program in light of the provisions enshrined in the new law.

An early scientific report on acromegaly: solving an intriguing endocrinological (c)old case?

We present and discuss a late-nineteenth century clinical case described by Professor Taruffi in a scientific paper titled "Scheletro con prosopoectasia e tredici vertebre dorsali" (Skeleton with prosopoectasia and thirteen thoracic vertebrae). Taruffi could not explain the disproportionate skeletal and visceral growth, and the case could therefore be considered an unrecognized case of acromegaly. The anatomopathological specimens and the wax model cited in the paper are currently hosted at the "Luigi Cattaneo" Anatomical Wax Collection of Bologna University; however, some inaccuracies and uncertainties as to their attribution to the same case have remained to this day. The skeletal remains were examined macroscopically to investigate any structural abnormalities and pathological changes. In addition, thanks to archival, museum inventory and literature research, we documented the systematic relationship between the paper and the samples and were able to ascribe the abnormally dilated dried stomach, currently displayed in a different showcase, to the same case. This is, to our knowledge, the first case of acromegaly in the history of medical literature which also includes a visceral specimen. As far as we know, there are no reports of the occurrence of severe gastromegaly in patients with acromegaly. In view of this rare association and, to date, endocrinological research, we hypothesize a further pathogenic mechanism by which acromegaly could have induced this massive dilatation. Taruffi's work represents an immensely valuable scientific/artistic heritage and is still cited in contemporary endocrinological literature, demonstrating its relevant contribution to the historical evolution of the disease through the nineteenth and twentieth centuries.

Enhancing radiosensitivity of melanoma cells through very high dose rate pulses released by a plasma focus device.

Radiation therapy is a useful and standard tumor treatment strategy. Despite recent advances in delivery of ionizing radiation, survival rates for some cancer patients are still low because of recurrence and radioresistance. This is why many novel approaches have been explored to improve radiotherapy outcome. Some strategies are focused on enhancement of accuracy in ionizing radiation delivery and on the generation of greater radiation beams, for example with a higher dose rate. In the present study we proposed an in vitro research of the biological effects of very high dose rate beam on SK-Mel28 and A375, two radioresistant human melanoma cell lines. The beam was delivered by a pulsed plasma device, a "Mather type" Plasma Focus for medical applications. We hypothesized that this pulsed X-rays generator is significantly more effective to impair melanoma cells survival compared to conventional X-ray tube. Very high dose rate treatments were able to reduce clonogenic efficiency of SK-Mel28 and A375 more than the X-ray tube and to induce a greater, less easy-to-repair DNA double-strand breaks. Very little is known about biological consequences of such dose rate. Our characterization is preliminary but is the first step toward future clinical considerations.

Marine bisindole alkaloid: A potential apoptotic inducer in human cancer cells.

Marine organisms such as corals, sponges and tunicates produce active molecules which could represent a valid starting point for new drug development processes. Among the various structural classes, the attention has been focused on 2,2-bis(6-bromo-3-indolyl) ethylamine, a marine alkaloid which showed a good anticancer activity against several tumor cell lines. Here, for the first time, the mechanisms of action of 2,2-bis(6-bromo-3-indolyl) ethylamine have been evaluated in a U937 tumor cell model. Morpho-functional and molecular analyses, highlighting its preferred signaling pathway, demonstrated that apoptosis is the major death response induced by this marine compund. Chromatin condensation, micronuclei formation, blebbing and in situ DNA fragmentation, occurring through caspase activation (extrinsic and intrinsic pathways), were observed. In particular, the bisindole alkaloid induces a mitochondrial involvement in apoptosis machinery activation with Blc-2/Bcl-x down-regulation and Bax up-regulation. These findings demonstrated that 2,2-bis(6-bromo-3-indolyl) ethylamine alkaloid-induced apoptosis is regulated by the Bcl-2 protein family upstream of caspase activation.

Phosphatidylinositol 3-kinase inhibition potentiates glucocorticoid response in B-cell acute lymphoblastic leukemia.

Despite remarkable progress in polychemotherapy protocols, pediatric B-cell acute lymphoblastic leukemia (B-ALL) remains fatal in around 20% of cases. Hence, novel targeted therapies are needed for patients with poor prognosis. Glucocorticoids (GCs) are drugs commonly administrated for B-ALL treatment. Activation of the phosphatidylinositol 3-kinase (PI3K)/Akt/mammalian target of rapamycin signaling pathway is frequently observed in B-ALL and contributes to GC-resistance. Here, we analyzed for the first time to our knowledge, the therapeutic potential of pan and isoform-selective PI3K p110 inhibitors, alone or combined with dexamethasone (DEX), in B-ALL leukemia cell lines and patient samples. We found that a pan PI3K p110 inhibitor displayed the most powerful cytotoxic effects in B-ALL cells, by inducing cell cycle arrest and apoptosis. Both a pan PI3K p110 inhibitor and a dual γ/δ PI3K p110 inhibitor sensitized B-ALL cells to DEX by restoring nuclear translocation of the GC receptor and counteracted stroma-induced DEX-resistance. Finally, gene expression analysis documented that, on one hand the combination consisting of a pan PI3K p110 inhibitor and DEX strengthened the DEX-induced up- or down-regulation of several genes involved in apoptosis, while on the other, it rescued the effects of genes that might be involved in GC-resistance. Overall, our findings strongly suggest that PI3K p110 inhibition could be a promising strategy for treating B-ALL patients by improving GC therapeutic effects and/or overcoming GC-resistance.

Improving nelarabine efficacy in T cell acute lymphoblastic leukemia by targeting aberrant PI3K/AKT/mTOR signaling pathway.

BACKGROUND: Although in recent years, the introduction of novel chemotherapy protocols has improved the outcome of T cell acute lymphoblastic leukemia (T-ALL) patients, refractory and/or relapsing disease remains a foremost concern. In this context, a major contribution was provided by the introduction of the nucleoside analog nelarabine, approved for salvage treatment of T-ALL patients with refractory/relapsed disease. However, nelarabine could induce a life-threatening, dose-dependent neurotoxicity. To improve nelarabine efficacy, we have analyzed its molecular targets, testing selective inhibitors of such targets in combination with nelarabine. METHODS: The effectiveness of nelarabine as single agent or in combination with PI3K, Bcl2, and MEK inhibitors was evaluated on human T-ALL cell lines and primary T-ALL refractory/relapsed lymphoblasts. The efficacy of signal modulators in terms of cytotoxicity, induction of apoptosis, and changes in gene and protein expression was assessed by flow cytometry, western blotting, and quantitative real-time PCR in T-ALL settings. RESULTS: Treatment with nelarabine as a single agent identified two groups of T-ALL cell lines, one sensitive and one resistant to the drug. Whereas sensitive T-ALL cells showed a significant increase of apoptosis and a strong down-modulation of PI3K signaling, resistant T-ALL cells showed a hyperactivation of AKT and MEK/ERK1/2 signaling pathways, not caused by differences in the expression of nelarabine transporters or metabolic activators. We then studied the combination of nelarabine with the PI3K inhibitors (both pan and dual γ/δ inhibitors), with the Bcl2 specific inhibitor ABT199, and with the MEK inhibitor trametinib on both T-ALL cell lines and patient samples at relapse, which displayed constitutive activation of PI3K signaling and resistance to nelarabine alone. The combination with the pan PI3K inhibitor ZSTK-474 was the most effective in inhibiting the growth of T-ALL cells and was synergistic in decreasing cell survival and inducing apoptosis in nelarabine-resistant T-ALL cells. The drug combination caused AKT dephosphorylation and a downregulation of Bcl2, while nelarabine alone induced an increase in p-AKT and Bcl2 signaling in the resistant T-ALL cells and relapsed patient samples. CONCLUSIONS: These findings indicate that nelarabine in combination with PI3K inhibitors may be a promising therapeutic strategy for the treatment of T-ALL relapsed patients.

Synergistic cytotoxic effects of bortezomib and CK2 inhibitor CX-4945 in acute lymphoblastic leukemia: turning off the prosurvival ER chaperone BIP/Grp78 and turning on the pro-apoptotic NF-κB.

The proteasome inhibitor bortezomib is a new targeted treatment option for refractory or relapsed acute lymphoblastic leukemia (ALL) patients. However, a limited efficacy of bortezomib alone has been reported. A terminal pro-apoptotic endoplasmic reticulum (ER) stress/unfolded protein response (UPR) is one of the several mechanisms of bortezomib-induced apoptosis. Recently, it has been documented that UPR disruption could be considered a selective anti-leukemia therapy. CX-4945, a potent casein kinase (CK) 2 inhibitor, has been found to induce apoptotic cell death in T-ALL preclinical models, via perturbation of ER/UPR pathway. In this study, we analyzed in T- and B-ALL preclinical settings, the molecular mechanisms of synergistic apoptotic effects observed after bortezomib/CX-4945 combined treatment. We demonstrated that, adding CX-4945 after bortezomib treatment, prevented leukemic cells from engaging a functional UPR in order to buffer the bortezomib-mediated proteotoxic stress in ER lumen. We documented that the combined treatment decreased pro-survival ER chaperon BIP/Grp78 expression, via reduction of chaperoning activity of Hsp90. Bortezomib/CX-4945 treatment inhibited NF-κB signaling in T-ALL cell lines and primary cells from T-ALL patients, but, intriguingly, in B-ALL cells the drug combination activated NF-κB p65 pro-apoptotic functions. In fact in B-cells, the combined treatment induced p65-HDAC1 association with consequent repression of the anti-apoptotic target genes, Bcl-xL and XIAP. Exposure to NEMO (IKKγ)-binding domain inhibitor peptide reduced the cytotoxic effects of bortezomib/CX-4945 treatment. Overall, our findings demonstrated that CK2 inhibition could be useful in combination with bortezomib as a novel therapeutic strategy in both T- and B-ALL.

Advances in understanding the acute lymphoblastic leukemia bone marrow microenvironment: From biology to therapeutic targeting.

The bone marrow (BM) microenvironment regulates the properties of healthy hematopoietic stem cells (HSCs) localized in specific niches. Two distinct microenvironmental niches have been identified in the BM, the "osteoblastic (endosteal)" and "vascular" niches. Nevertheless, these niches provide sanctuaries where subsets of leukemic cells escape chemotherapy-induced death and acquire a drug-resistant phenotype. Moreover, it is emerging that leukemia cells are able to remodel the BM niches into malignant niches which better support neoplastic cell survival and proliferation. This review focuses on the cellular and molecular biology of microenvironment/leukemia interactions in acute lymphoblastic leukemia (ALL) of both B- and T-cell lineage. We shall also highlight the emerging role of exosomes/microvesicles as efficient messengers for cell-to-cell communication in leukemia settings. Studies on the interactions between the BM microenvironment and ALL cells have led to the discovery of potential therapeutic targets which include cytokines/chemokines and their receptors, adhesion molecules, signal transduction pathways, and hypoxia-related proteins. The complex interplays between leukemic cells and BM microenvironment components provide a rationale for innovative, molecularly targeted therapies, designed to improve ALL patient outcome. A better understanding of the contribution of the BM microenvironment to the process of leukemogenesis and leukemia persistence after initial remission, may provide new targets that will allow destruction of leukemia cells without adversely affecting healthy HSCs. This article is part of a Special Issue entitled: Tumor Microenvironment Regulation of Cancer Cell Survival, Metastasis,Inflammation, and Immune Surveillance edited by Peter Ruvolo and Gregg L. Semenza.

PI3K pan-inhibition impairs more efficiently proliferation and survival of T-cell acute lymphoblastic leukemia cell lines when compared to isoform-selective PI3K inhibitors.

Class I phosphatidylinositol 3-kinases (PI3Ks) are frequently activated in T-cell acute lymphoblastic leukemia (T-ALL), mainly due to the loss of PTEN function. Therefore, targeting PI3Ks is a promising innovative approach for T-ALL treatment, however at present no definitive evidence indicated which is the better therapeutic strategy between pan or selective isoform inhibition, as all the four catalytic subunits might participate in leukemogenesis. Here, we demonstrated that in both PTEN deleted and PTEN non deleted T-ALL cell lines, PI3K pan-inhibition exerted the highest cytotoxic effects when compared to both selective isoform inhibition or dual p110γ/δ inhibition. Intriguingly, the dual p110γ/δ inhibitor IPI-145 was effective in Loucy cells, which are representative of early T-precursor (ETP)-ALL, a T-ALL subtype associated with a poor outcome. PTEN gene deletion did not confer a peculiar reliance of T-ALL cells on PI3K activity for their proliferation/survival, as PTEN was inactivated in PTEN non deleted cells, due to posttranslational mechanisms. PI3K pan-inhibition suppressed Akt activation and induced caspase-independent apoptosis. We further demonstrated that in some T-ALL cell lines, autophagy could exert a protective role against PI3K inhibition. Our findings strongly support clinical application of class I PI3K pan-inhibitors in T-ALL treatment, with the possible exception of ETP-ALL cases.

Therapeutic potential of targeting mTOR in T-cell acute lymphoblastic leukemia (review).

T-cell acute lymphoblastic leukemia (T-ALL) is a heterogeneous neoplastic disorder of immature hematopoietic precursors committed to the T-cell lineage. T-ALL comprises about 15% of pediatric and 25% of adult ALL cases. Even if the prognosis of T-ALL has improved especially in the childhood due to the use of new intensified treatment protocols, the outcome of relapsed patients who are resistant to conventional chemotherapeutic drugs or who relapse is still poor. For this reason, there is a need for novel and less toxic targeted therapies against signaling pathways aberrantly activated in T-ALL, such as the phosphoinositide 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR). Small molecules designed to target key components of this signaling axis have proven their efficacy both in vitro and in vivo in pre-clinical settings of T-ALL. In particular, different classes of mTOR inhibitors have been disclosed by pharmaceutical companies, and they are currently being tested in clinical trials for treating T-ALL patients. One of the most promising approaches for the treatment of T-ALL seems to be the combination of mTOR inhibitors with traditional chemotherapeutic agents. This could lead to a lower drug dosage that may circumvent the systemic side effects of chemotherapeutics. In this review, we focus on the different classes of mTOR inhibitors that will possibly have an impact on the therapeutic arsenal we have at our disposal against T-ALL.

Targeting signaling pathways in T-cell acute lymphoblastic leukemia initiating cells.

Leukemia initiating cells (LICs) represent a reservoir that is believed to drive relapse and resistance to chemotherapy in blood malignant disorders. T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive neoplastic disorder of immature hematopoietic precursors committed to the T-cell lineage. T-ALL comprises about 15% of pediatric and 25% of adult ALL cases and is prone to early relapse. Although the prognosis of T-ALL has improved especially in children due to the use of new intensified treatment protocols, the outcome of relapsed T-ALL cases is still poor. Putative LICs have been identified also in T-ALL. LICs are mostly quiescent and for this reason highly resistant to chemotherapy. Therefore, they evade treatment and give rise to disease relapse. At present great interest surrounds the development of targeted therapies against signaling networks aberrantly activated in LICs and important for their survival and drug-resistance. Both the Notch1 pathway and the phosphoinositide 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) network are involved in T-ALL LIC survival and drug-resistance and could be targeted by small molecules. Thus, Notch1 and PI3K/Akt/mTOR inhibitors are currently being developed for clinical use either as single agents or in combination with conventional chemotherapy for T-ALL patient treatment. In this review, we summarize the existing knowledge of the relevance of Notch1 and PI3K/Akt/mTOR signaling in T-ALL LICs and we examine the rationale for targeting these key signal transduction networks by means of selective pharmacological inhibitors.

Osteobiography of a 19th century elderly woman with pertrochanteric fracture and osteoporosis: a multidisciplinary approach.

In this paper the osteobiography of an elderly woman recovered from a cemetery tomb where she was buried in 1850, affected by hip fracture and osteoporosis, is described. The overall anthropological characteristics of the individual have been investigated. Macroscopic, radiographic, tomographic, microscopic, and chemical and structural examinations have been performed to give a detailed account of the condition of the skeleton. A non-union pertrochanteric fracture not surgically treated and probably due to senile osteoporosis was diagnosed. The consequences of the fracture to the bones show that this individual likely survived several years following the injury. The osseous features we describe (remodelled bone at the fracture site, asymmetry of entheseal changes likely related to the particular walking pattern of the individual) may be useful in personal identification of skeletons of legal interest. Regarding the recognition of osteoporosis in unearthed skeletons, our study underlines that the cortical thickness, microscopic features, degree of crystallinity and Ca/P ratio represent more useful elements than the mean bone density, mineral/matrix ratio and mineral maturity, which are more sensitive to diagenetic changes that affect the mineral phase post-mortem.

Differential proteomic and phenotypic behaviour of papillary and anaplastic thyroid cell lines.

Thyroid carcinomas account for a minority of all malignant tumours but, after those of the gonads, they represent the most common forms of endocrine cancers. They include several types, among which the papillary thyroid cancer (PTC) and the anaplastic thyroid cancer (ATC) are the best known. The two hystotypes display significant biological and clinical differences: PTC is a well differentiated form of tumour with a high incidence and a good prognosis, while the ATC is less frequent but represents one of the most aggressive endocrine tumours with morphological features of an undifferentiated type. To date, as far as we know, no conclusive studies, useful to design arrays of molecular markers, have been published illustrating the phenotypic and proteomic differences between these two tumours. The aim of this work was to perform a comparative analysis of two thyroid cancer cell lines, derived respectively from papillary (BCPAP) and anaplastic (8505C) thyroid carcinomas. The comparative analysis included cell behaviour assays and proteomic analysis by 2D-PAGE and mass spectrometry. The results have highlighted a new proteomic signature for the anaplastic carcinoma-derived cells, consistent with their high proliferation rate, motility propensity and metabolic shift, in relation to the well-differentiated PTC cells.

A combination of temsirolimus, an allosteric mTOR inhibitor, with clofarabine as a new therapeutic option for patients with acute myeloid leukemia.

Signaling through the phosphatidylinositol 3-kinase (PI3K) pathway and its downstream effectors, Akt and mechanistic target of rapamycin (mTOR), is aberrantly activated in acute myeloid leukemia (AML) patients, where it contributes to leukemic cell proliferation, survival, and drug-resistance. Thus, inhibiting mTOR signaling in AML blasts could enhance their sensitivity to cytotoxic agents. Preclinical data also suggest that allosteric mTOR inhibition with rapamycin impaired leukemia initiating cells (LICs) function. In this study, we assessed the therapeutic potential of a combination consisting of temsirolimus [an allosteric mTOR complex 1 (mTORC1) inhibitor] with clofarabine, a nucleoside analogue with potent inhibitory effects on both ribonucleotide reductase and DNA polymerase. The drug combination (CLO-TOR) displayed synergistic cytotoxic effects against a panel of AML cell lines and primary cells from AML patients. Treatment with CLO-TOR induced a G0/G1-phase cell cycle arrest, apoptosis, and autophagy. CLO-TOR was pro-apoptotic in an AML patient blast subset (CD34⁺/CD38⁻/CD123⁺), which is enriched in putative leukemia initiating cells (LICs). In summary, the CLO-TOR combination could represent a novel valuable treatment for AML patients, also in light of its efficacy against LICs.

Early healing events around titanium implant devices with different surface microtopography: a pilot study in an in vivo rabbit model.

In the present pilot study, the authors morphologically investigated sandblasted, acid-etched surfaces (SLA) at very early experimental times. The tested devices were titanium plate-like implants with flattened wide lateral sides and jagged narrow sides. Because of these implant shape and placement site, the device gained a firm mechanical stability but the largest portion of the implant surface lacked direct contact with host bone and faced a wide peri-implant space rich in marrow tissue, intentionally created in order to study the interfacial interaction between metal surface and biological microenvironment. The insertion of titanium devices into the proximal tibia elicited a sequence of healing events. Newly formed bone proceeded through an early distance osteogenesis, common to both surfaces, and a delayed contact osteogenesis which seemed to follow different patterns at the two surfaces. In fact, SLA devices showed a more osteoconductive behavior retaining a less dense blood clot, which might be earlier and more easily replaced, and leading to a surface-conditioning layer which promotes osteogenic cell differentiation and appositional new bone deposition at the titanium surface. This model system is expected to provide a starting point for further investigations which clarify the early cellular and biomolecular events occurring at the metal surface.

Dental implant thread pitch and its influence on the osseointegration process: an in vivo comparison study.

PURPOSE: The design of an implant plays a fundamental role in the osseointegration process, particularly in low-density bone. It has been postulated that design features that maximize the surface area available for contact may improve mechanical anchorage and primary stability in cancellous bone. Two different implant profiles were compared to evaluate the influence of thread pitch on the osseointegration process in bone of low density and limited height. MATERIALS AND METHODS: "Narrow-pitch" implants (NP) with a 0.5-mm pitch and "wide-pitch" implants (WP) with a 1.7-mm pitch were tested for osseointegration after 0 days and 4 and 8 weeks in a sheep iliac crest model. The two different implants were analyzed with biologic and biomechanical tests. RESULTS: The present findings showed that initial mechanical anchorage and subsequent early endosseous integration in low-density bone could be improved by a reduction of thread pitch. The greater surface area gained by decreasing thread pitch increased bone-implant contact and primary stability from the time of implant placement. This better performance of the NP profile could be appreciated even at an early healing time when the subsequent biologic integration was enhanced not only in terms of a higher quantity of newly deposited bone but also more regular and mature geometric distribution of bone tissue at the interface. CONCLUSION: These results confirmed that, when primary stability is a concern, as in cancellous bone, increasing the implant surface area by using implants with smaller pitch might be beneficial.

Correlative microscopy of bone in implant osteointegration studies.

Routine morphological analyses usually include investigations by light microscopy (LM), scanning electron microscopy (SEM), and transmission electron microscopy (TEM). Each of these techniques provides specific information on tissue morphology and all the obtained results are then combined to give an in-depth morphological overview of the examined sample. The limitations of this traditional comparative microscopy lie in the fact that each technique requires a different experimental sample, so that many specimens are necessary and the combined results come from different samples. The present study describes a technical procedure of correlative microscopy, which allows us to examine the same bone section first by LM and then, after appropriate processing, by SEM or TEM. Thanks to the possibility of analyzing the same undecalcified bone sections both by LM and SEM, the approach described in the present study allows us to make very accurate evaluations of old/new bone morphology at the bone-implant interface.

Collagen structure of tendon relates to function.

A tendon is a tough band of fibrous connective tissue that connects muscle to bone, designed to transmit forces and withstand tension during muscle contraction. Tendon may be surrounded by different structures: 1) fibrous sheaths or retinaculae; 2) reflection pulleys; 3) synovial sheaths; 4) peritendon sheaths; 5) tendon bursae. Tendons contain a) few cells, mostly represented by tenoblasts along with endothelial cells and some chondrocytes; b) proteoglycans (PGs), mainly decorin and hyaluronan, and c) collagen, mostly type I. Tendon is a good example of a high ordered extracellular matrix in which collagen molecules assemble into filamentous collagen fibrils (formed by microfibrils) which aggregate to form collagen fibers, the main structural components. It represents a multihierarchical structure as it contains collagen molecules arranged in fibrils then grouped in fibril bundles, fascicles and fiber bundles that are almost parallel to the long axis of the tendon, named as primary, secondary and tertiary bundles. Collagen fibrils in tendons show prevalently large diameter, a D-period of about 67 nm and appear built of collagen molecules lying at a slight angle (< 5 degrees). Under polarized light microscopy the collagen fiber bundles appear crimped with alternative dark and light transverse bands. In recent studies tendon crimps observed via SEM and TEM show that the single collagen fibrils suddenly changing their direction contain knots. These knots of collagen fibrils inside each tendon crimp have been termed "fibrillar crimps", and even if they show different aspects they all may fulfil the same functional role. As integral component of musculoskeletal system, the tendon acts to transmit muscle forces to the skeletal system. There is no complete understanding of the mechanisms in transmitting/absorbing tensional forces within the tendon; however it seems likely that a flattening of tendon crimps may occur at a first stage of tendon stretching. Increasing stretching, other transmission mechanisms such as an interfibrillar coupling via PGs linkages and a molecular gliding within the fibrils structure may be involved.

FBS suppresses TGF-beta1-induced chondrogenesis in synoviocyte pellet cultures while dexamethasone and dynamic stimuli are beneficial.

In vitro cartilage tissue engineering culture systems benefit from a fine balance of biochemical and mechanical components to maintain the chondrocyte phenotype. This balance, however, can be disrupted by using typical methods for cultivating chondrogenic cells in medium supplemented with fetal bovine serum (FBS) and growth factors. Our goal was to determine the effects of fluid-dynamic stimuli, fetal bovine serum and dexamethasone on the chondrogenesis of 14-day synoviocyte pellet cultures in the presence of TGF-beta1. We employed a pellet culture system that provides a highly cellular three-dimensional structure that permits differentiation and extracellular matrix synthesis. Our results indicated that FBS inhibited glycosaminoglycan (GAG) and type II collagen production. Interestingly, the effect of dynamic stimuli was modulated by the presence of FBS; mixed serum-free cultures had increased GAG production, whereas mixed cultures with 10% FBS exhibited less GAG production compared with their static counterparts, possibly due to pronounced suppressive effects of FBS via increased transport. Dexamethasone addition during the first week of culture resulted in enhanced extracellular matrix production and increased cellularity. Moreover, the presence of 10% FBS in addition to ITS(+) and TGF-beta1 did not significantly increase cell proliferation compared with serum-free medium. These results indicate the importance of a comprehensive analysis of growth conditions for each cell culture system.