[Use of cultured human autologous bone marrow stem cells in repair of a rotator cuff tear: preliminary results of a safety study]. / Pouzití kultivovaných lidských autologních kmenových bunek kostní drene pri rekonstrukci ruptury rotátorové manzety - studie bezpecnosti metody, predbezné výsledky.

PURPOSE OF THE STUDY Rotator cuff tears are one of the most frequent shoulder disorders which are often associated with pain and interfere with proper arm function. In order to evaluate the safety and effectiveness of using cultured human autologous mesenchymal stem cells (MSC) applied to the suture site during arthroscopic repair of a rotator cuff tear, a prospective clinical study was designed and started recently at the authors' department. Its primary goal was to evaluate the safety of using cultured human MSCs, the secondary goal then was to study a therapeutic effect of their application. Preliminary results of the study on a limited number of patients are presented here. MATERIAL AND METHODS Ten patients who met the indication criteria for arthroscopic repair of a rotator cuff tear were included in the study. In addition, they also had to meet inclusion and lack exclusion criteria. According to the protocol, their bone marrow was harvested at 3 to 4 weeks before surgery. Subsequently, an arthroscopic repair of the rotator cuff tear was performed and an suspension of cultured MSCs was applied to the suture site at the end of the procedure. The isolation of MSCs from bone marrow and their cultivation was carried out by the company Bioinova, Ltd. The patients were followed up at 6 weeks and 3 and 6 months post-operatively. Their clinical assessment included physical examination of the shoulder, pain intensity evaluation according to the visual analogue scale (VAS), and subjective questionnaires for Constant and University of California (UCLA) scores. All patients underwent MRI examination at 6 post-operative months to evaluate the quality of rotator cuff reconstruction. The findings were compared with the pre-operative results. RESULTS A final evaluation was made in eight patients of 10. Two patients were excluded from the study because their exclusion criteria were fulfilled. The evaluated patients showed significantly better clinical outcomes as early as 6 weeks after surgery; also all pre-operative scores were improved at 3 and 6 months. The average values at 6 months post-operatively were: 0 points for the VAS score, 32 for the UCLA score and 84 for the Constant score. The MRI findings at 6 months after surgery showed fully healed and well-integrated tissue of the rotator cuff tendon attachment in all eight patients. No adverse effects of therapy were recorded during the follow-up period. DISCUSSION The use of autologous stem cells and growth factors in the treatment of tendons, muscles and cartilage is currently the topic of many experimental studies on animal models. Its utilisation in human clinical trials has been reported only marginally; the relevant studies have so far used only suspensions of non-cultured mononuclear cells. Our study, although on a smallsize patient group, provides evidence that human cultured autologous MSCs can safely be used for tissue repair in the indications mentioned above. CONCLUSIONS Our preliminary short-term results show that using human cultured autologous MSCs in the treatment of rotator cuff tears is safe. However, further research is needed, particularly with regard to the effectiveness of the method. Key words: rotator cuff tear, arthroscopic repair, mesenchymal stem cells, tendon, cell therapy.

Stem cells regenerative properties on new rat spinal fusion model.

Stem cells biology is one of the most frequent topic of physiological research of today. Spinal fusion represents common bone biology challenge. It is the indicator of osteoinduction and new bone formation on ectopic model. The purpose of this study was to establish a simple model of spinal fusion based on a rat model including verification of the possible use of titanium microplates with hydroxyapatite scaffold combined with human bone marrow-derived mesenchymal stem cells (MSCs). Spinous processes of two adjacent vertebrae were fixed in 15 Wistar rats. The space between bony vertebral arches and spinous processes was either filled with augmentation material only and covered with a resorbable collagen membrane (Group 1), or filled with augmentation material loaded with 5 × 106 MSCs and covered with a resorbable collagen membrane (Group 2). The rats were sacrificed 8 weeks after the surgery. Histology, histomorphometry and micro-CT were performed. The new model of interspinous fusion was safe, easy, inexpensive, with zero mortality. We did not detect any substantial pathological changes or tumor formation after graft implantation. We observed a nonsignificant effect on the formation of new bone tissue between Group 1 and Group 2. In the group with MSCs (Group 2) we described minor inflamatory response which indicates the imunomodulational and antiinflamatory role of MSCs. In conclusion, this new model proved to be easy to use in small animals like rats.

Both autologous bone marrow mononuclear cell and peripheral blood progenitor cell therapies similarly improve ischaemia in patients with diabetic foot in comparison with control treatment.

BACKGROUND: The aim of our study was to compare the effect of bone marrow mononuclear cell and peripheral blood progenitor cell therapies in patients with diabetic foot disease and critical limb ischaemia unresponsive to revascularization with conservative therapy. METHODS: Twenty-eight patients with diabetic foot disease (17 treated by bone marrow cells and 11 by peripheral blood cell) were included into an active group and 22 patients into a control group without cell treatment. Transcutaneous oxygen pressure and rate of major amputation, as the main outcome measures, were compared between bone marrow cells, peripheral blood cell and control groups over 6 months; both cell therapy methods were also compared by the characteristics of cell suspensions. Possible adverse events were evaluated by changes of serum levels of angiogenic cytokines and retinal fundoscopic examination. RESULTS: The transcutaneous oxygen pressure increased significantly (p < 0.05) compared with baseline in both active groups after 6 months, with no significant differences between bone marrow cells and peripheral blood cell groups; however, no change of transcutaneous oxygen pressure in the control group was observed. The rate of major amputation by 6 months was significantly lower in the active cell therapy group compared with that in the control group (11.1% vs. 50%, p = 0.0032), with no difference between bone marrow cells and peripheral blood cell. A number of injected CD34+ cells and serum levels of angiogenic cytokines after treatment did not significantly differ between bone marrow cells and peripheral blood cell. CONCLUSIONS: Our study showed a superior benefit of bone marrow cells and peripheral blood cell treatments of critical limb ischaemia in patients with diabetic foot disease when compared with conservative therapy. There was no difference between both cell therapy groups, and no patient demonstrated signs of systemic vasculogenesis.

Experimental reconstruction of the injured spinal cord.

Injury to the spinal cord, with its pathological sequelae, results in a permanent neurological deficit. With currently available tools at hand, there is very little that clinicians can do to treat such a condition with the view of helping patients with spinal cord injury (SCI). On the other hand, in the last 20 years experimental research has brought new insights into the pathophysiology of spinal cord injury; we can divide the time course into 3 phases: primary injury (the time of traumatic impact and the period immediately afterwards), the secondary phase (cell death, inflammation, ischemia), and the chronic phase (scarring, demyelination, cyst formation). Increased knowledge about the pathophysiology of SCI can stimulate the development of new therapeutic modalities and approaches, which may be feasible in the future in clinical practice. Some of the most promising experimental therapies include: neurotrophic factors, enzymes and antibodies against inhibitory molecules (such as Nogo), activated macrophages, stem cells and bridging scaffolds. Their common goal is to reconstitute the damaged tissue in order to recover the lost function. In the current review, we focus on some of the recent developments in experimental SCI research.

[Treatment of critical limb ischemia and diabetic foot disease by the use of autologous stem cells]. / Terapie kritické koncetinové ischemie u pacientu se syndromem diabetické nohy pomocí autologních kmenových bunek.

AIM: The aim of our study was to assess safety and effectiveness of therapy of critical limb ischaemia by autologous stem cells and evaluation of potential adverse events. METHODS: Fourteen patients were included into the study (11 men, 3 women, mean age 61.9 +/- 9.6 years, mean diabetes duration 23.5 +/- 11.1 years, mean glycated hemoglobin 6 +/- 1%). Eight patients were treated by bone marrow stromal cells, 6 patients by peripheral blood progenitor cells after stimulation by filgrastim. The suspension of stem cells was then applied into the muscles of ischemic limbs. We evaluated transcutaneous oxygen tension (TcPO2), subjective pain sensation assessed by Visual Analog Scale (VAS) and wound healing. RESULTS: TcPO2 significantly increased in all patients from 10 +/- 8.7 mm Hg before the treatment to 39.4 +/- 9.5 mm Hg after 6 months (p = 0.0005) after stem cell therapy. We also observed significant area defect reduction and pain decrease during the follow-up period. Median of area defect was reduced from 4.3 (0.7 - 31.7) before the treatment to 0.06 (0 - 0.5) cm2 after 6 months from the treatment (p = 0.0078). Decrease in rest pain was observed in all patients, mean VAS decreased from 5.3 +/- 1.8 to 1.1 +/- 1.3 after 6 months (p = 0.002). CONCLUSION: Our study suggests that stem cell therapy of diabetic foot disease is an effective therapeutic option with no adverse events for patients with severe peripheral arterial disease. This treatment leads to increase of transcutaneous oxygen tension, improves wound healing and decreases the rest pain.

The diffusion parameters of the extracellular space are altered in focal cortical dysplasias.

Most hypotheses concerning the mechanisms underlying seizure activity in focal cortical dysplasia (FCD) are based on alterations in synaptic transmission and glial dysfunction. However, neurons may also communicate by extrasynaptic transmission, which was recently found to affect epileptiform activity under experimental conditions and which is mediated by the diffusion of neuroactive substances in the extracellular space (ECS). The ECS diffusion parameters were therefore determined using the real-time iontophoretic method in human neocortical tissue samples obtained from surgically treated epileptic patients. The obtained values of the extracellular space volume fraction and tortuosity were then correlated with the histologicaly assessed type of cortical malformation (FCD type I or II). While the extracellular volume remained unchanged (FCD I) or larger (FCD II) than in normal/control tissue, tortuosity was significantly increased in both types of dysplasia, indicating the presence of additional diffusion barriers and compromised diffusion, which might be another factor contributing to the epileptogenicity of FCD.

[Comparison of chondrogenic differentiation of adipose tissue-derived mesenchymal stem cells with cultured chondrocytes and bone marrow mesenchymal stem cells]. / Výsledky srovnání chondrogenní diferenciace mezenchymálních kmenových bunek získaných z tukové tkáne s kultivovanými chondrocyty a mezenchymálními kmenovými bunkami z kostní drene

PURPOSE OF THE STUDY: Many congenital and acquired disorders as well as sequelae of injury are associated with articular cartilage degeneration, which adversely affects the patient's quality of life. The currently used cell therapy with cultured chondrocytes has its disadvantages due to a process of de-differentiation of chondrocytes during cultivation. We believe that the mesenchymal stem cell therapy offers a new treatment options. MATERIAL AND METHODS: The adult mesenchymal stem cells (MSCs) for chondrocyte differentiation are usually obtained from bone marrow mesenchymal stem cells (BMSCs). In this study these cells were compared with mesenchymal stem cells derived from adipose tissue (AMSCs). The aim of the study was to verify the ability of human BMSCs and AMSCs to differentiate into chondrocytes in vitro in the presence or absence of transforming growth factor beta (TGF-ß1). Human BMSCs and AMSCs were collected from healthy donors during orthopaedic surgeries, in vitro cultured in three passages to obtain the required quantity of cells. A pellet culture system was used for chondrocyte differentiation. RESULTS: At 21 days of cultivation, cell aggregates grown in the chondrogenic medium were larger than those cultured in the control medium. Both the BMSCs and AMSCs pellet cultures showed spontaneous chondrogenesis. Histological staining with haematoxylin and eosin and Masson's trichrome stains, as well as immunohistochemical staining to detect type II collagen revealed no apparent differences between the pellet cultures with TGF-ß1 presence and those without it. The real-time RT-PCR detected expression of the type II collagen gene in all tested cultures. In the BMSCs pellet culture only, TGF-ß1 presence resulted in a decrease in type I collagen mRNA levels and in an increase in type II collagen mRNA values. DISCUSSION: Our results showed an in vitro chondrogenic potential of mature human mesenchymal stem cells derived from both bone marrow and adipose tissue. In agreement with the relevant literature data, we suggest that both cell types have an equal prospect for use in cartilage tissue engineering.

Macroporous hydrogels based on 2-hydroxyethyl methacrylate. Part 6: 3D hydrogels with positive and negative surface charges and polyelectrolyte complexes in spinal cord injury repair.

Macroporous hydrogels are artificial biomaterials commonly used in tissue engineering, including central nervous system (CNS) repair. Their physical properties may be modified to improve their adhesion properties and promote tissue regeneration. We implanted four types of hydrogels based on 2-hydroxyethyl methacrylate (HEMA) with different surface charges inside a spinal cord hemisection cavity at the Th8 level in rats. The spinal cords were processed 1 and 6 months after implantation and histologically evaluated. Connective tissue deposition was most abundant in the hydrogels with positively-charged functional groups. Axonal regeneration was promoted in hydrogels carrying charged functional groups; hydrogels with positively charged functional groups showed increased axonal ingrowth into the central parts of the implant. Few astrocytes grew into the hydrogels. Our study shows that HEMA-based hydrogels carrying charged functional groups improve axonal ingrowth inside the implants compared to implants without any charge. Further, positively charged functional groups promote connective tissue infiltration and extended axonal regeneration inside a hydrogel bridge.

Measuring diffusion parameters in the brain: comparing the real-time iontophoretic method and diffusion-weighted magnetic resonance.

The extracellular space (ECS) diffusion parameters influence the movement of ions, neuroactive substances, hormones and metabolites in the nervous tissue. They also affect extrasynaptic transmission, a mode of signal transmission dependent solely on diffusion. This review compares in detail two methods for studying diffusion in the brain: the real-time iontophoretic tetramethylammonium method for ECS volume fraction and tortuosity measurements and diffusion weighted-magnetic resonance imaging for measuring the apparent diffusion coefficient of water. The results obtained using both methods under physiological conditions (post-natal development, ageing) or in pathologies (brain injury, ischaemia) and their similarities and differences are discussed.

Rapid but not slow spinal cord compression elicits neurogenic pulmonary edema in the rat.

The development of neurogenic pulmonary edema (NPE) can be elicited by an immediate epidural balloon compression of the thoracic spinal cord. To evaluate whether a slower balloon inflation could prevent NPE development, we examined the extent of NPE in animals lesioned with a rapid (5 microl - 5 microl - 5 microl) or slow rate (3 microl - 2 microl - 2 microl - 2 microl - 2 microl - 2 microl - 2 microl) of balloon inflation. These groups were compared with the NPE model (immediate inflation to 15 microl) and with healthy controls. Slow balloon inflation prevented NPE development, whereas the pulmonary index and histology revealed a massive pulmonary edema in the group with a rapid rate of balloon inflation. Pulmonary edema was preceded by a considerable decrease in heart rate during the inflation procedure. Moreover, rapid inflation of balloon in spinal channel to either 5 microl or 10 microl did not cause NPE. Thus, a slow rate of balloon inflation in the thoracic epidural space prevents the development of neurogenic pulmonary edema, most likely due to the better adaptation of the organism to acute circulatory changes (rapid elevation of systemic blood pressure accompanied by profound heart rate reduction) during the longer balloon inflation period. It should be noted that spinal cord transection at the same level did not cause neurogenic pulmonary edema.

Biocompatible hydrogels in spinal cord injury repair.

Spinal cord injury results in a permanent neurological deficit due to tissue damage. Such a lesion is a barrier for "communication" between the brain and peripheral tissues, effectors as well as receptors. One of the primary goals of tissue engineering is to bridge the spinal cord injury and re-establish the damaged connections. Hydrogels are biocompatible implants used in spinal cord injury repair. They can create a permissive environment and bridge the lesion cavities by providing a scaffold for the regeneration of neurons and their axons, glia and other tissue elements. The advantage of using artificial materials is the possibility to modify their physical and chemical properties in order to develop the best implant suitable for spinal cord injury repair. As a result, several types of hydrogels have been tested in experimental studies so far. We review our work that has been done during the last 5 years with various types of hydrogels and their applications in experimental spinal cord injury repair.

Extracellular space diffusion and extrasynaptic transmission.

The diffusion of neuroactive substances in the extracellular space (ECS) plays an important role in short- and long-distance communication between nerve cells and is the underlying mechanism of extrasynaptic (volume) transmission. The diffusion properties of the ECS are described by three parameters: 1. ECS volume fraction alpha (alpha=ECS volume/total tissue volume), 2. tortuosity lambda (lambda2=free/apparent diffusion coefficient), reflecting the presence of diffusion barriers represented by, e.g., fine neuronal and glial processes or extracellular matrix molecules and 3. nonspecific uptake k'. These diffusion parameters differ in various brain regions, and diffusion in the CNS is therefore inhomogeneous. Moreover, diffusion barriers may channel the migration of molecules in the ECS, so that diffusion is facilitated in a certain direction, i.e. diffusion in certain brain regions is anisotropic. Changes in the diffusion parameters have been found in many physiological and pathological states in which cell swelling, glial remodeling and extracellular matrix changes are key factors influencing diffusion. Changes in ECS volume, tortuosity and anisotropy significantly affect the accumulation and diffusion of neuroactive substances in the CNS and thus extrasynaptic transmission, neuron-glia communication, transmitter "spillover" and synaptic cross-talk as well as cell migration, drug delivery and treatment.

Mechanisms of neurogenic pulmonary edema development.

Neurogenic pulmonary edema is a life-threatening complication, known for almost 100 years, but its etiopathogenesis is still not completely understood. This review summarizes current knowledge about the etiology and pathophysiology of neurogenic pulmonary edema. The roles of systemic sympathetic discharge, central nervous system trigger zones, intracranial pressure, inflammation and anesthesia in the etiopathogenesis of neurogenic pulmonary edema are considered in detail. The management of the patient and experimental models of neurogenic pulmonary edema are also discussed.

Low degree of anesthesia increases the risk of neurogenic pulmonary edema development.

Neurogenic pulmonary edema is an acute life-threatening complication following central nervous system injury. The exact pathogenic mechanism leading to its development is still unclear. We introduce a new hypothesis that high levels of anesthesia might protect the organism against the development of neurogenic pulmonary edema due to a more pronounced inhibition of the hypothalamic, brainstem and spinal vasoactive sympathetic centers. On the basis of a more pronounced neuronal inhibition of the vasoactive centers, a severe sympathetic discharge does not occur and neurogenic pulmonary edema does not develop. In contrast, an insufficient anesthesia level is not able to inhibit the sympathetic nervous system during an injury of the central nervous system and thus neurogenic pulmonary edema develops. During experiments with central nervous system injury, low-anesthesia-induced neurogenic pulmonary edema might negatively influence the overall recovery of the animal. More importantly, during a neurosurgical intervention, insufficient anesthesia might similarly lead to neurogenic pulmonary edema development in operated patients. Our hypothesis indicates the necessity of precisely monitoring of the level anesthesia during experimental manipulations of the central nervous system in animals or neurosurgical interventions in humans.

Lesion evolution after gamma knife irradiation observed by magnetic resonance imaging.

PURPOSE: Our study is focused on the magnetic resonance imaging (MRI) observation of lesion development and hippocampus related functional impairments in rats after irradiation with a Leksell Gamma knife (LGK). MATERIALS AND METHODS: We exposed 32 three-month-old Long-Evans rats to various radiation doses (25 Gy, 50 Gy or 75 Gy). The rats were scanned by a 4.7 T magnetic resonance (MR) spectrometer at several timepoints (1 - 18 months) after irradiation. The lesion size was evaluated by manual segmentation; the animals were behaviorally tested in a Morris water maze and examined histologically. RESULTS: We found that a dose of 25 Gy induced no edema, necrosis or behavioral change. The response of the rats to higher doses was not uniform; the first occurrence of lesions in the rat brains irradiated with 50 and 75 Gy was detected six months post-irradiation. Functional impairment correlated well with the lesion size and histology. CONCLUSIONS: Rat brains showed the development of expanding delayed lesions after 50 or 75 Gy doses from the LGK during the first year after irradiation.

Migration, fate and in vivo imaging of adult stem cells in the CNS.

Adult stem cells have been intensively studied for their potential use in cell therapies for neurodegenerative diseases, ischemia and traumatic injuries. One of the most promising cell sources for autologous cell transplantation is bone marrow, containing a heterogenous cell population that can be roughly divided into hematopoietic stem and progenitor cells and mesenchymal stem cells (MSCs). MSCs are multipotent progenitor cells that, in the case of severe tissue ischemia or damage, can be attracted to the lesion site, where they can secrete bioactive molecules, either naturally or through genetic engineering. They can also serve as vehicles for delivering therapeutic agents. Mobilized from the marrow, sorted or expanded in culture, MSCs can be delivered to the damaged site by direct or systemic application. In addition, MSCs can be labeled with superparamagnetic nanoparticles that allow in vivo cell imaging. Magnetic resonance imaging (MRI) is thus a suitable method for in vivo cell tracking of transplanted cells in the host organism. This review will focus on cell labeling for MRI and the use of MSCs in experimental and clinical studies for the treatment of brain and spinal cord injuries.

Macroporous hydrogels based on 2-hydroxyethyl methacrylate. Part 4: growth of rat bone marrow stromal cells in three-dimensional hydrogels with positive and negative surface charges and in polyelectrolyte complexes.

The growth of bone marrow stromal cells was assessed in vitro in macroporous hydrogels based on 2-hydro- xyethyl methacrylate (HEMA) copolymers with different electric charges. Copolymers of HEMA with sodium methacrylate (MA(-)) carried a negative electric charge, copolymers of HEMA with [2-(methacryloyloxy)ethyl] trimethylammonium chloride (MOETA(-)) carried a positive electric charge and terpolymers of HEMA, MA(-) and MOETA(+) carried both, positive and negative electric charges. The charges in the polyelectrolyte complexes were shielded by counter-ions. The hydrogels had similar porosities, based on a comparison of their diffusion parameters for small cations as measured by the real-time tetramethylammonium iontophoretic method of diffusion analysis. The cell growth was studied in the peripheral and central regions of the hydrogels at 2 hours and 2, 7, 14 and 28 days after cell seeding. Image analysis revealed the highest cellular density in the HEMA-MOETA(+) copolymers; most of the cells were present in the peripheral region of the hydrogels. A lower density of cells but no difference between the peripheral and central regions was observed in the HEMA-MA(-) copolymers and in polyelectrolyte complexes. This study showed that positively charged functional groups promote the adhesion of cells.

Temporal profile of ultrastructural changes in cortical neurones after a compression lesion.

We studied the occurrence of apoptosis and secondary delayed cell death at various time points in the penumbra zone, which is the target for therapeutic intervention after stroke. A compression lesion was induced in the right sensory motor cortex of rat brains. At 0.5, 1, 3, 6, 12, 24, 48 and 72 h after lesioning, motor functions were evaluated by behavioral tests, and cortical layers IV and V were examined by electron microscopy. Behavioral recovery was observed at 48 h after lesioning. At 0.5-1 h in the lesioned area, the neuropil was expanded and contained affected cells. Apoptotic cells were found between 0.5-72 h, and at 12 h, 47.3 % of the total cell number was apoptotic cells. On the contralateral side, cells showed an enlarged endoplasmic reticulum at 3 h, indicating secondary delayed cell death. Our results show that a compression lesion is a useful model for studying ultrastructural changes in injured cells. The lesion results in the penumbra zone with apoptotic cell death between 0.5-72 h. As secondary delayed cell death occurred on the contralateral side at three hours after lesioning might be the time period during which injured, but still viable, neurons can be targets for acute treatment.

Macroporous hydrogels based on 2-hydroxyethyl methacrylate. Part II. Copolymers with positive and negative charges, polyelectrolyte complexes.

Crosslinked macroporous hydrogels based on 2-hydroxyethyl methacrylate (HEMA)-[2-(methacryloyloxy)ethyl]trimethylammonium chloride (MOETACl) copolymer, HEMA-MOETACl-methacrylic acid (MA) terpolymer, and on a polyelectrolyte complex of HEMA-MA copolymer with poly(MOETACl) were prepared. All the hydrogels were prepared in the presence of fractionated sodium chloride particles. The hydrogels were characterized by the number of pores and the total volume of all pores in unit volume, the average volume and the average diameter of single pore. Morphology of the hydrogels was investigated by confocal and scanning electron microscopy. The hydrogels based on polyelectrolyte complexes were also characterized by chemical composition. Homogeneous (non-porous) hydrogels with the same composition as macroporous hydrogels were prepared and characterized by their biocompatibility.

[The role of the extracellular space in biology of glial brain tumors]. / Význam mezibunecnéeho prostoru v biologii gliových nádoru mozku.

The size, geometry and composition of the extracellular space (ECS) play an important role in influencing the biological behavior of primary brain tumors. Experiments employing the real-time TMA iontophoretic method to determine the size and geometry of the ECS, by monitoring the diffusion of TMA ions in the ECS, revealed a dramatic increase in ECS size in brain neoplasms when compared with that of unaffected brain cortex. Further, the increase of ECS volume in tumors was shown to correlate with increasing proliferative activity and increasing cellularity of astrocytomas. The increase in ECS size was surprisingly accompanied by a significant increase in diffusion barriers, slowing the diffusion of molecules in the ECS of tumors. In low-grade tumors, diffusion is hindered by the presence of a dense net of tumor cell processes. In high-grade gliomas, in which the cellular processes are shortened with reduced branching, the increase in diffusion barriers is caused by the overproduction of specific components of the extracellular matrix (ECM) by the tumor cells, mainly tenascin. The ECM glycoproteins produced represent a substrate for the subsequent adhesion and migration of tumor cells through the enlarged ECS. However, they might also critically reduce the diffusion of therapeutics into the tumor. The presence of tenascin in the ECS of a neoplasm correlates significantly with the increased malignancy of the tumor and a poor clinical outcome of the disease, thus making the immunohistochemical detection of tenascin diagnostically useful as a prognostic marker and a marker of aggressive biological behavior of tumors.