Endovascular Recanalization of Tandem Internal Carotid Occlusions Using the Balloon-assisted Tracking Technique.

PURPOSE: Tandem occlusive lesions are responsible for up to 20% of acute ischemic stroke cases and are associated with poor prognosis if complete recanalization cannot be achieved. Endovascular recanalization might be challenging due to difficulties in the safe passage of the occluded plaque at the origin of the internal carotid artery (ICA). The balloon-assisted tracking technique (BAT), where a partially deflated balloon is exposed out of the catheter tip to facilitate its passage through stenosed or spastic arterial segments was introduced by interventional cardiologists and the applicability of the technique has been recently proposed in the field of neurointervention as well. Here we describe our experience using the BAT technique in the endovascular recanalization of tandem occlusive lesions. METHODS: Procedures were performed from June 2013 to December 2020 in a single center. Baseline clinical and imaging data, procedural and follow-up details and clinical outcomes were retrospectively collected. RESULTS: In this study 107 patients, median age 66 years, median admission NIHSS 14 and median ASPECTS 8 were included. Successful recanalization of the ICA using the BAT technique was achieved in 100 (93%) and successful intracranial revascularization in 88 (82%) patients. There were no complications attributable to the BAT technique. Intraprocedural complications occurred in 9 (8%) patients. Emergent stenting was performed in 40 (37%) at the end of the procedure. Postprocedural adverse events (intracerebral hemorrhage [ICH], malignant infarction) occurred in 6 (5%) patients. Good clinical outcome at 3 months (modified Rankin scale [mRS] 0-2) was 54 (50%) and mortality 26 (24%). Delayed stent placement during follow-up occurred in 21 cases. CONCLUSION: Application of BAT technique in tandem occlusions appears feasible, safe, and efficient. Further evaluation of this technique is awaited.

ITIH5 shows tumor suppressive properties in cervical cancer cells grown as multicellular tumor spheroids.

Cervical cancer (CC) arises from premalignant cervical intraepithelial neoplasia (CIN) induced by a persistent infection with human papillomaviruses. The multi-stepwise disease progression is driven by genetic and epigenetic alterations. Our previous studies demonstrated a clear downregulation of inter-α-trypsin-inhibitor-heavy chain 5 (ITIH5) at mRNA and protein levels in CC compared to CIN2/3 and normal cervical tissue. Initial in vitro functional analyses revealed a suppressive effect of ITIH5 on relevant mechanisms for cancer progression in conventional two dimensional (2D) cell culture model systems. Based on these studies, we aimed to investigate the functional relevance of ITIH5 in multicellular tumor spheroid (MCTS) models, which resemble in vivo tumors more closely. We successfully established CC cell line-derived MCTS using the hanging-drop technique. ITIH5 was ectopically overexpressed in HeLa and SiHa cells and its functional relevance was investigated under three dimensional (3D) culture conditions. We found that ITIH5 re-expression significantly suppressed tumor spheroid growth and spheroid invasiveness of both HeLa and SiHa spheroids. Immunohistochemical (IHC) analyses revealed a significant reduction in Ki-67 cell proliferation index and CAIX-positive areas indicative for hypoxia and acidification. Furthermore, we observed an increase in cPARP-positive cells suggesting a higher rate of apoptosis upon ITIH5 overexpression. An effect of ITIH5 expression on the susceptibility of cervical MCTS towards cytostatic drug treatment was not observed. Collectively, these data uncover pronounced anti-proliferative effects of ITIH5 under 3D cell culture conditions and provide further functional evidence that the downregulation of ITIH5 expression during cervical carcinogenesis could support cancer development.

Successful management of zinc phosphide poisoning-a Hungarian case.

BACKGROUND: Zinc phosphide (ZnP) is the basic component of several insecticides easily accessible worldwide. Intentional or accidental intoxication may lead to severe complications and multiple organ failure, resulting in high mortality. No known antidote is currently available. The iron-chelation and the antioxidative effects are well-known features of alpha-lipoic acid (ALA), although its use in the treatment of ZnP poisoning has not been documented previously. We describe the case of a patient with serious ZnP poisoning with multiple organ failure, where ALA was also included in the patient's supportive therapy. CASE PRESENTATION: A 65-year-old man ingested 125 g of Arvalin® (containing 5 g ZnP) and presented to the Emergency Department, with respiratory insufficiency and decreased consciousness. He developed hypokalemia, hypocalcemia, low white blood cell count, elevated C-reactive protein level, mixed acidosis, hepatic and kidney damage, thickening of the jejunal wall, and lung atelectasis, which served as a basis for the ensuing bacterial pneumonia. Antibiotics and adequate supportive therapy were provided. Laboratory tests indicated liver damage (slightly increased liver enzymes, low pseudocholinesterase levels; 706 U/L on day 2), possibly caused by the patient's chronic alcoholism or the ZnP poison itself, therefore, hepatoprotective agents, ALA (Thiogamma Turbo-Set®) with N-acetylcysteine were administered for six consecutive days. Pseudocholinesterase values increased sixfold until the end of the second week of care. Fifteen days after admission, the patient was relocated to the department of psychiatry with stable vital functions, clear consciousness, declining inflammatory markers, and improved liver function. He was discharged 1 month later, fully recovered. CONCLUSIONS: Our case is the first documented voluntary and severe ZnP poisoning in Hungary. Our patient developed multiple organ failure and atelectasis, possibly resulting in the observed respiratory infection. The development of bacterial pneumonia highlighted the dangers of phosphine-induced atelectasis. The use of ALA in our patient's case, as an antioxidant and agent for metal chelation, suggested that this agent could be a promising tool in the prevention and treatment of ZnP-induced hepatic damage.

Breast cancer, placenta and pregnancy.

BACKGROUND: Breast cancer is one of the most frequently diagnosed malignancies during pregnancy. Tumours often present characteristics of high malignancy and are hormone receptor negative/HER2 positive or triple negative. In general, pregnancy, including the postpartum period, is associated with a transiently increased risk of developing breast cancer but followed by a long-lasting protective period. Placental metastases are very rare and, thus far, breast cancer metastases in the foetal compartment have not been described. To discuss these apparently contradictory observations, this narrative review resumes immunological and hormonal alterations during pregnancy potentially affecting breast cancer risk as well as tumour growth and behaviour. OBSERVATIONS: Upregulation of breast cancer-associated genes involved in immunological and reproductive processes has been observed in parous women and is potentially responsible for a transiently increased risk in pregnancy. In contrast, maternal immunisation and immunoglobulin production against antigens expressed on trophoblast cells, such as specific glycosylation patterns of mucin-1 or RCAS1-associated truncated glycans, seem to prevent breast cancer development in later years. Animal and human studies indicate that T cells are involved in these processes. Several placenta-derived factors, especially kisspeptin, have direct anti-tumour effects. The pregnancy-related increase of estrogen, progesterone, and other hormones influence growth and characteristics of breast cancer while the role of further placenta-secreted factors is still controversially discussed. CONCLUSION: Several factors and cells are involved in altered breast cancer risk during and after pregnancy and have potential for developing novel treatment strategies in future.

Human serum alters cell culture behavior and improves spheroid formation in comparison to fetal bovine serum.

BACKGROUND: The use of fetal bovine serum (FBS) as growth supplement for human cell and tissue culture is widely spread in basic research as well as in clinical approaches, although several limitations must be considered, such as unstable composition and availability, biosafety and ethical aspects. Regarding interspecies differences, xenogeneic growth factors may evoke incompatibilities and non-desired interactions with human cells resulting in imprecise outcome of human-relevant data. METHODS: In this study the functionality of human serum (HS) has been investigated in comparison to FBS by assessing proliferation, migration and invasion of the human cervical cancer cell lines SiHa and HeLa. The effects of both sera on spheroid formation were analyzed microscopically. RESULTS: Both, FBS and HS, stimulate cell proliferation and migration similarly, whereas HS significantly enhanced cell invasion. The spheroid formation assay revealed remarkable differences between both sera, especially for SiHa cells. While in FBS supplemented medium cells only formed loose aggregates, HS induced regularly shaped spheroids under all tested conditions. CONCLUSION: We were able to demonstrate that HS and FBS differently influence behavior of cells in culture which may have an impact on experimental results, especially in 3D cultures.

Uterine natural killer cells in patients with idiopathic recurrent miscarriage.

PROBLEM: Uterine natural killer (uNK) cells are major players during implantation and early pregnancy. The aim of our study was to analyze uNK cell concentration in the endometrium of idiopathic recurrent miscarriage (iRM) patients and fertile controls. METHOD OF STUDY: Out of n=130 couples with ≥3 consecutive, clinical RM screened according to a standardized diagnostic protocol, n=58 patients with iRM were identified. Endometrial biopsies were investigated in patients and n=17 fertile women (controls) via immunohistochemistry. RESULTS: Compared to controls, the concentration of uNK cells was significantly higher in iRM patients (257±212 vs. 148±73 uNK cells/mm², P=.04). IRM patients showed a higher prevalence of >300 uNK cells/mm² than controls (34.5% vs. 5.9%, P=.02). In 88% of controls and 62% of iRM patients, uNK cells were detected within the range of 40-300/mm². CONCLUSION: Idiopathic recurrent miscarriage patients showed higher uNK cell levels than controls supporting the possible impact of uNK cells in the pathophysiology of miscarriage. Our cutoff levels might help to select RM patients which may benefit from immunomodulatory treatment.

Generation of Multicellular Breast Cancer Tumor Spheroids: Comparison of Different Protocols.

Multicellular tumor spheroids are widely used models in tumor research. Because of their three dimensional organization they can simulate avascular tumor areas comprising proliferative and necrotic cells. Nonetheless, protocols for spheroid generation are still inconsistent. Therefore, in this study the breast cancer cell lines MCF-7, MDA-MB-231 and SK-BR-3 have been used to compare different spheroid generation models including hanging drop, liquid overlay and suspension culture techniques, each under several conditions. Experimental approaches differed in cell numbers (400-10,000), media and additives (25 % methocel, 25 % methocel plus 1 % Matrigel, 3.5 % Matrigel). In total, 42 different experimental setups have been tested. Generation of spheroids was evaluated by light microscopy and the structural composition was assessed immunohistochemically by means of Ki-67, cleaved poly (ADP-ribose) polymerase (cPARP) and mucin-1 (MUC-1) expression. Although the tested cell lines diverged widely in their capacity of forming spheroids we recommend hanging drops supplemented with 25 % methocel as the most reliable and efficient method with regard to success of generation of uniform spheroids, costs, experimental complexity and time expenditure in the different cell lines. MCF-7 cells formed spheroids under almost all analyzed conditions, and MDA-MB-231 cells under only one protocol (liquid overlay technique, 3.5 % Matrigel), while SK-BR-3 did not under neither condition. Therefore, we outline specific methods and recommend the use of adapted and standardized spheroid generation protocols for each cell line.

Anti-inflammatory effects of limb ischaemic preconditioning are mediated by sensory nerve activation in rats.

We have shown that ischaemic preconditioning ameliorates both the local periosteal and the systemic leukocyte activation evoked by limb ischaemia-reperfusion. We hypothesized that the activation of chemosensitive afferent nerves by transient ischaemia contributes to the protective mechanisms of ischaemic preconditioning via a calcitonin gene-related peptide (CGRP)-dependent mechanism. In Sprague-Dawley rats, 60-min complete limb ischaemia was followed by 180 min of reperfusion. In further experiments, the CGRP analogue hCGRP (0.3 µg kg(-1)) or ischaemic preconditioning (2 × 10-min ischaemia/10-min reperfusion) was applied prior to the ischaemia-reperfusion insult. Ischaemic preconditioning was performed in three subgroups in which animals received the CGRP receptor antagonist CGRP(8-37) (30 µg kg(-1) h(-1)), the chemosensitive afferent nerve inactivator resiniferatoxin (3 × 15 µg kg(-1), sc), or vehicle. The effects of CGRP(8-37) and resiniferatoxin on ischaemia-reperfusion without ischaemic preconditioning were also evaluated. In the tibial periosteum of rats, intravital fluorescence microscopy and immunohistochemistry revealed significant attenuations of ischaemia-reperfusion-induced post-ischaemic leukocyte-endothelial interactions (rolling and adherence in the postcapillary venules) and tissue intracellular adhesion molecule expression following ischaemic preconditioning or hCGRP administration. Administration of CGRP(8-37) or pretreatment of animals with resiniferatoxin reversed the anti-inflammatory effects of limb ischaemic preconditioning, but failed to affect the microcirculatory consequences of ischaemia-reperfusion without ischaemic preconditioning. The results suggest that activation of the chemo- (capsaicin-) sensitive afferent nerves is involved in the mechanisms of microcirculatory anti-inflammatory protection provided by limb ischaemic preconditioning. Controlled activation of chemosensitive C-fibres or the CGRP receptors by the induction of ischaemic preconditioning or other means may furnish therapeutic benefit by ameliorating the periosteal microcirculatory consequences of tourniquet ischaemia.

20-Hydroxyecdysone increases fiber size in a muscle-specific fashion in rat.

20-Hydroxyecdysone (20E) is an ecdysteroid hormone that regulates moulting in insects. Interestingly, 20E is also found most abundantly in plant species and has anabolic effects in vertebrates, i.e. increasing muscle size without androgen influence. The effect of 20E on slow and fast fiber types of skeletal muscle has not been reported yet. Here we present that 20E affects the size (cross-sectional area, CSA) of the different fiber types in a muscle-specific manner. The effect on fiber size was modified by the distance from the site of the treatment and the presence of a regenerating soleus muscle in the animal. Besides the fiber size, 20E also increased the myonuclear number in the fibers of normal and regenerating muscles, suggesting the activation of satellite cells. According to our results 20E may provide an alternative for substitution of anabolic-androgenic steroids in therapeutic treatments against muscle atrophy.