Analysis of Connexin 43 and Spermine Co-localisation in Glioblastomas.

Gap junctions are channels between adjacent cells, contributing to the unhindered exchange of metabolites, second messengers, nucleotides, and other molecules. The functional status of gap junctions in brain tumours is underinvestigated. One avenue of research focuses on exploring the expression of polyamines and their co-localisation with the Connexin 43 (Cx43) in the growth zones of glioblastoma multiforme (GBM). The aim of this work was to analyse the expression of Cx43 and spermine in human GBM to reveal their roles in neuro-oncogenesis. Human GBM sample sections were used for the immunochemistry [glial fibrillary acidic protein (GFAP), Cx43, and spermine], confocal laser scanning microscopy, and electron immunohistochemistry. Immunofluorescent analysis revealed that the more extensive processes of GBM cells exhibit GFAP. All GBM samples (n = 10) exhibited positive Cx43 signals in the form of variously sized dots and lines. Cx43 formed dotted lines around cell bodies with segmented transformed nuclei, which were also present in the gliovascular complexes. Furthermore, spermine was overexpressed in all tumour samples (cytoplasm and large and thin tumour processes), including the areas of Cx43 localisation. Merging the Cx43 and spermine signals showed co-expression in the same regions: the membranes of individual cells and individual points on processes in the tumour tissue. Therefore, we established the staining of the co-localisation of Cx43 and the polyamine spermine within glioblastoma, revealing that tumour processes housing the polyamine indeed form gap junctions, suggesting their potential joint interaction. This finding indicates that glioma cells can integrate into the surrounding neural networks, potentially serving as a mechanism to release glycolysis products, relying on gap junction activity facilitated by spermine. Cx43 exhibits sensitivity to polyamines, which play a role in opening gap junctional channels. Furthermore, polyamines have been observed to eliminate the blockades caused by hydrogen ions and calcium, which is crucial for cellular physiology.

Confocal Laser and Electron Microscopic Investigation of Gap Junctions in Anaplastic Astrocytomas.

Connexin 43 (Cx43) is a multifunction protein that forms gap junction channels and hemichannels and is suggested to play an essential role in oxygen-glucose deprivation, induced via neuroinflammation during astrocytoma expansion into healthy tissue. To prove this assumption we studied connexin 43 localisation and ultrastructure of gap junctions in samples of malignant brain tumour (anaplastic astrocytomas grade III). For confocal laser microscopy, vibratome sections of tumour fragments were incubated in a mixture of primary antibodies to connexin 43 and glial fibrillary acidic protein (GFAP), then in a mixture of secondary antibodies conjugated with a fluorescent label. After the immunofluorescence study, sections were washed in phosphate buffer, additionally postfixed with 1% OsO4 solution, dehydrated and embedded in epoxy resin by a plane-parallel method. Ultra-thin sections obtained from these samples were contrasted with uranyl acetate and lead citrate and viewed under a Jem 1011 electron microscope. Confocal laser examination detected a positive reaction to Cx43 in the form of point fluorescence. These points were of various sizes. Most of them were localised around or at the intersection of small processes containing GFAP. Electron microscopy of the tumour samples containing the most significant number of Cx43 revealed single and closely spaced gap junctions with a typical ultrastructure on the processes and bodies of tumour cells. Sequential analysis in the fields of view revealed 62 gap junctions in the area of 100 µm2. Numerous gap junctions in anaplastic astrocytomas revealed in our study may indicate electrotonic and metabolic transmission between glioma cells, possibly promoting its progression.

How I do it: decompressive hemicraniectomy supplemented with resection of the temporal pole and tentoriotomy for malignant ischemic infarction in the territory supplied by the middle cerebral artery.

Malignant ischemic infarction in the territory supplied by the middle cerebral artery is an extremely severe form of ischemic stroke associated with development of massive uncontrollable postischemic edema of the affected cerebral hemisphere; the end result of which is development of transtentorial herniation and death. METHOD: The surgical technique of performance of decompressive hemicraniectomy involves removal of an extensive bone flap in the fronto-temporo-parieto-occipital zone with resection of the temporal squama and of the greater wing of the sphenoid bone to visualize the level of entrance of the middle meningeal artery to the cranial cavity, which, in its turn, allows resection of the upright margin of the middle cranial fossa. Decompressive hemicraniectomy is supplemented with resection of the temporal pole and tentoriotomy. CONCLUSION: Performance of decompressive hemicraniectomy in combination with resection of the resection of the temporal pole and tentoriotomy is an effective surgical method of treatment of malignant ischemic stroke in the territory supplied by the middle cerebral artery, capable of reducing the lethality rate during the postoperative period.