Jürgen A. Hampl

Integration of Functional Data (PET) into Brain Surgery Planning and Neuronavigation

There are distinct advantages of including functional data from PET into surgery planning and neuronavigation: tumor delineation and localization of functional areas. Preoperative intervention planning in brain tumours poses many additional challenges if the goal is to include functional data from PET for neuronavigation: co-registration of multimodal image data (MRI and PET) is conceptually the most demanding task. However, we find that associated problems of visualization, data transfer, documentation and, in general terms, quality assurance are often underestimated. We address some of these problems, including visual inspection of co-registration accuracy, preparation of image data, DICOM capabilities and “electronic documentation” [1].

Hypertrophy of the lumbar ligamentum flavum is associated with inflammation-related TGF-β expression

BackgroundDespite the significance of hypertrophy of the ligamentum flavum (HLF) in the disease progress of neurogenic claudication, the cellular mechanisms underlying the gradual fibrotic thickening of the ligamentum flavum remain poorly understood. The aim of our study was to get insight into the contribution of inflammatory mechanisms to the development of hypertrophy.MethodsSpecimens of hypertrophied ligamenta flava were obtained at surgery from 20 patients with acquired lumbar osteoligamentous spinal canal stenosis from the central part of the ligament. Paraffin sections were stained with hematoxylin and eosin and Elastica van Gieson to evaluate extracellular matrix architecture, and immunohistochemistry was performed to characterize the inflammatory reaction and the sources of transforming growth factor beta (TGF-β) expression. Sections of normal ligamenta flava obtained from corresponding anatomical sites and stained in parallel served as a control.ResultsHLF was characterized by a considerable distortion of the elastic matrix and fibrotic transformation by extracellular collagen deposition. All specimens showed highly inflammatory cellular infiltrates confined to regions exhibiting marked degeneration of the elastic matrix composed mainly of macrophages, scattered T lymphocytes, and neovascularization, thus representing a chronic inflammation. Surprisingly, macrophages as well as vascular endothelial cells but not fibroblasts showed a strong expression of TGF-β, a strong inducer of extracellular collagen deposition.ConclusionsMacrophages were identified as a major cellular source of TGF-β in advanced HLF and may perpetuate further hypertrophy. This finding suggests that modulating the immune response locally or systemically could prove to be effective for impeding the disease progress.

Potentiated Gene Delivery to Tumors Using Herpes Simplex Virus/Epstein-Barr Virus/RV Tribrid Amplicon Vectors

The development and use of gene transfer techniques creates an opportunity to achieve better treatment modalities for numerous disease entities. Promising results for treatment in tumor cells in culture and in small animal models have been reported. Nevertheless, the lack of widespread vector distribution throughout tumor tissue is one of the current limitations for successful clinical application of gene therapy paradigms. The use of migratory tumor cells themselves as vector delivery vehicles may allow wider vector distribution in tumors. In addition, continuous release of retrovirus vectors on-site could generate a high local virion concentration over an extended time period with consequent increases in transduction efficiency. In this paper, we present in culture and in vivo data of a herpes simplex virus-Epstein-Barr virus hybrid amplicon vector containing retrovirus vector components (tribrid vector) that allows conversion of tumor cells into retroviral producer cells. With this method, we were able to achieve a local fourfold amplification of stable transgene expression in tumors. The application of this system, which can integrate a transgene cassette into tumors with therapeutic bystander effects, could increase the local amplification effect to a level of clinical relevance.

Generation of stable retrovirus packaging cell lines after transduction with herpes simplex virus hybrid amplicon vectors.

A number of properties have relegated the use of Moloney murine leukemia virus (Mo‐MLV)‐based retrovirus vectors primarily to ex vivo protocols. Direct implantation of retrovirus producer cells can bypass some of the limitations, and in situ vector production may result in a large number of gene transfer events. However, the fibroblast nature of most retrovirus packaging cells does not provide for an effective distribution of vector producing foci in vivo, especially in the brain. Effective development of new retrovirus producer cells with enhanced biologic properties may require the testing of a large number of different cell types, and a quick and efficient method to generate them is needed.

Developmental potential of the murine embryonic stem cells transplanted into the healthy rat brain--novel insights into tumorigenesis.

Although engraftment of undifferentiated pluripotent embryonic stem cells (ESCs) into the injured central nervous system (CNS) may lead to targeted cell replacement of lost/damaged cells, sustained proliferative activity combined with uncontrolled differentiation of implanted cells presents a risk of tumor formation. As tumorigenic potential is thought to be associated with pluripotency of embryonic stem cells, pre-differentiation may circumvent this problem. Recently, it has been demonstrated that tumorigenesis occurs despite pre-differentiation if the neural precursor cells are implanted into the brain of a homologous animal (e.g., mouse to mouse). However, xenotransplantation (e.g., mouse to rat) without pre-differentiation, lead to the development of healthy neuronal cells, in absence of tumor formation, suggesting that tumor-suppressive effects of host tissue on engrafted ESCs may play a role in transplant tumorigenesis. We critically investigated tumorigenesis and possible mechanisms of anticipated tumor-suppressive effect under conditions analogous to previously published studies. Xenotransplantation of D-3 murine ESCs into uninjured adult rat brains lacking any preliminary inflammatory potential was found to lead to tumor formation in 5 out of 8 of animals within 2 weeks postimplantation. Tumor-suppressive effects, reflected by Erdo et. al could possibly be ascribed to immunomodulatory activity of macrophages scavenging the tumorigenic fraction of the implanted cells. The importance of number of engrafted cells, implantation site and immunosuppressive effects are discussed as possible variables determining tumorigenic outcome after ESC transplantation.

Extract Derived from Rat Brains in the Acute Phase Following Traumatic Brain Injury Impairs Survival of Undifferentiated Stem Cells and Induces Rapid Differentiation of Surviving Cells

Dramatic cerebral responses following brain injury (TBI) comprise inflammation, cell death, and modulation of trophic factor release. These cerebral modulations might induce and /or attenuate acute neuronal damage. Here, we investigated the effect of tissue extract derived from healthy (HBE) or injured rat brain (TBE) on the differentiation of cultured embryonic stem cells in vitro. Rats were sacrificed at t = 45 minutes following lateral fluid-percussion injury and extracts of cerebral tissue were prepared from 4-6 healthy or injured rat brain hemispheres. Murine embryonic stem cells (CGR8) cultured in serum-free medium were then conditioned for a week with HBE or TBE. Omission of serum from the culture medium induced neural differentiation of CGR8 stem cells, as indicated by a significant time dependent down-regulation of oct-4 with a concomitant upregulation of nestin after 7 days. In parallel cell loss was observed that seemed to be largely due to apoptotic cell death. In TBE treated cells, on the other hand, a significant amplification of apoptotic cell death, enhancement of nestin and MAP2 expression and marked morphological changes such as axonal-like outgrowth was observed within 3 days of conditioning. Treatment of stem cells with HBE resulted in less pronounced neuronal differentiation processes. Axonal-like outgrowth was not observed. Our data suggest that during the early acute phase of traumatic injury the cerebral environment is disposed to detrimental as well as potent protective signals that seem to rapidly induce neurogenic processes.

Degeneration of cholinergic rat basal forebrain neurons after experimental subarachnoid hemorrhage.

OBJECTIVEThe reasons for neuropsychological deficits after subarachnoid hemorrhage (SAH) are fairly unknown. Cholinergic basal forebrain (BFB) neurons are essential for attention, memory, and emotion. We investigated possible changes in the cholinergic BFB and its hippocampal and neocortical terminals after experimental SAH. METHODSSAH was induced in 19 male Wistar rats by stereotactic injection of 150 μL of autologous blood into the prechiasmatic cistern. Five control animals received 150 μL of saline. Continuous monitoring of brain tissue oxygen tension, intracranial pressure, and cerebral perfusion pressure was performed. After 4 and 14 days, the BFB was analyzed for cholinergic and γ-aminobutyric acid-ergic cell counts. The number of cholinergic terminals in the hippocampus and neocortex was calculated by optical densitometry. RESULTSSAH resulted in a 20 to 30% decrease in cholinergic BFB neurons in the medial septum and diagonal band at 4 and 14 days. A similar decline in the density of hippocampal and neocortical cholinergic terminals was demonstrated. Animals treated with saline did not exhibit significant cholinergic cell loss, and γ-aminobutyric acid-ergic neurons appeared unaffected by the SAH. Courses of intracranial pressure and cerebral perfusion pressure did not differ between animals injected with blood and saline, but brain tissue oxygen tension decreased considerably and continued to stay below baseline in SAH, although it returned to normal values after saline injection. CONCLUSIONThe present study provides evidence for a decrease of cholinergic BFB neurons after SAH. The direct effect of blood in the basal cisterns seemed to result in an enduring tissue hypoxia as a significant mechanism for cholinergic degeneration.

Treatment strategies of space-occupying intradural metastases of the cauda equina of nonneurogenic origin

BackgroundCompressive intradural metastases of the cauda equina are a rare site of metastatic spread in systemic cancer. So far, only few reports have been published with conflicting statements concerning a surgical versus nonsurgical approach.MethodFive patients with symptomatic space-occupying intradural metastases of the cauda equina were analyzed retrospectively, focusing on the influence of surgical intervention on pain relief, neurological outcome and thus the patients’ quality of life.FindingsAt the time of diagnosis, all patients were in an advanced metastatic state. Surgical resection was the primary treatment in four patients and radiotherapy in one. Despite infiltration of the cauda rootlets, gross total tumour resection could be achieved in two of the four patients treated surgically. Functional outcome was beneficial in these patients with marked and immediate relief of pain and improvement of motor function even following incomplete tumour resection.ConclusionsSurgical treatment of compressive intradural metastases of the cauda equina seems to be feasible with low operative risk and with the potential benefit of an immediate relief of pain and improvement in motor function and thus an increase in quality of life.

Intracerebral Administration of Heat-Inactivated Staphylococcus Epidermidis Enhances Oncolysis and Prolongs Survival in a 9L Orthotopic Gliosarcoma Model

Background/Aims: The association between postoperative infection and prolonged survival in high-grade glioma is still a matter of debate. Previously we demonstrated that the intracerebral (i.c.) injection of heat-inactivated staphylococcal epitopes (HISE) resulted in a well-defined infux of immunocompetent cells across the blood-brain barrier. The present study investigated the potential antitumoral effect of HISE-immunostimulation in an experimental glioma model. Methods: Wistar rats were intracerebrally implanted with 9L gliosarcoma cells (n=6), 9L cells mixed with HISE (n=12), or phosphate buffered saline (n=4). Tumor growth was measured by serial magnetic resonance imaging (MRI). After death due to the tumor burden, the brains were histopathologically assessed for inflammation and oncolysis. A toxicity assay was performed to quantify potential impairment of HISE on tumor cell growth in vitro. Results: Animals treated by HISE showed a significant increase in average survival and even complete regression of an already established mass in one case. Naïve 9L gliosarcomas failed to recruit significant numbers of systemic immune cells. In contrast, concomitant intracerebral HISE inoculation lead to a oncolysis and a distinct peri- and intratumoral infiltration of macrophages, CD8 and CD4 co-expressing T-lymphocytes in two thirds of the tumor-bearing animals. The toxicity screening showed HISE-mediated oncolysis to be ineffective ex vivo. Conclusion: This study describes a novel approach for combatting malignant glioma using inactivated staphylococci as potent immunomodulators. Our results provide an outline for investigating the strategic potential of bacteria as emerging future therapeutics.

A novel experimental in vivo model of cerebral immunomodulation induced by inactivated Staphylococcus epidermidis

The genesis and appropriate treatment of neuroinflammation in various infectious and non-infectious disorders of the central nervous system is still a matter of debate. We introduce an alternative and simple experimental model for the investigation of the cellular inflammatory response to bacterial antigens by stereotactic intracerebral injection of heat-inactivated Staphylococcus epidermidis (HISE). HISE-injection resulted in well-circumscribed intraparenchymal deposits encompassed by an early micro- and astroglial response and a selective but sustained opening of the blood-brain barrier (BBB). After 24h, the HISE collections were densely infiltrated by granulocytes and few circumjacent macrophages that became the predominating immunocompetent cell type from day 4 on. CD8a+ lymphocytes peaked at day 4, whereas CD4+ and CD20+ lymphocytes increased gradually in number, developing a scattered infiltrate until day 17, indicating the initiation of an adaptive immune response. MHC class II presenting cells were abundantly recruited from day 1 and eventually shaped an increasingly dense accumulation within the lesion. Intracerebral HISE administration provides a controlled, highly reproducible and well defined influx of immunocompetent cells across the BBB leading to a distinct and condensed inflammatory reaction. The technique is straightforward, easily feasible and may significantly enable further investigations of the initiation, maintenance and therapeutic modulation of acute neuroinflammation.