Organotypic slice cultures of human glioblastoma reveal different susceptibilities to treatments

Background

Glioblastoma multiforme is the most common lethal brain tumor in human adults, with no major therapeutic breakthroughs in recent decades. Research is based mostly on human tumor cell lines deprived of their organotypic environment or inserted into immune-deficient animals required for graft survival. Here, we describe how glioblastoma specimens obtained from surgical biopsy material can be sectioned and transferred into cultures within minutes.

Methods

Slices were kept in 6-well plates, allowing direct observation, application of temozolomide, and irradiation. At the end of experiments, slice cultures were processed for histological analysis including hematoxylin-eosin staining, detection of proliferation (Ki67), apoptosis/cell death (cleaved caspase 3, propidium iodide), DNA double-strand breaks (γH2AX), and neural subpopulations. First clinical trials employed irradiation with the heavy ion carbon for the treatment of glioblastoma patients, but the biological effects and most effective dose regimens remain to be established. Therefore, we developed an approach to expose glioblastoma slice cultures to ¹²C and X-rays.

Results

We found preservation of the individual histopathology over at least 16 days. Treatments resulted in activation of caspase 3, inhibition of proliferation, and cell loss. Irradiation induced γH2AX. In line with clinical observations, individual tumors differed significantly in their susceptibility to temozolomide (0.4%–2.5% apoptosis and 1%–15% cell loss).

Conclusion

Glioblastoma multiforme slice cultures provide a unique tool to explore susceptibility of individual tumors for specific therapies including heavy ions, thus potentially allowing more personalized treatments plus exploration of mechanisms of (and strategies to overcome) tumor resistance.     

Transpetrosal approach: an anatomical study of temporal bone

Resection of the petrous temporal bone to various degrees provides different levels of access to lesions of the posterior fossa. However, regarding the numerous variations, precise distances of petrosal bone are not still clearly described. This may lead to serious complications during transpetrosal surgeries. Our objective was to evaluate different distances of temporal bone landmarks in order to assess their variations and the possible correlations between them. This anatomical study was performed on 60 temporal bones from 60 human cadavers in the years 2006 and 2007. All the bones contained an adequate portion of the petrous apex and attached fossa dura. Twelve landmarks were defined and 27 different distances were measured for each temporal bone using two-point caliper. Less variation was observed in the superoinferior diameter of horizontal carotid canal with the less coefficient of variation (CV) of 9.29; whereas, the most variation was detected in the inferior (axial) plane of posterior semicircular canal to superior plane of jugular bulb (CV = 57.65). There was a significant correlation between vertical intratemporal diameter of carotid in pyramidal direction, and superior–inferior diameter of horizontal carotid canal (r _Pearson = 0.500, P < 0.001). Other significant correlations were also found between other distances. The variations of different distances and landmarks were evaluated and many significant correlations were demonstrated between them which could potentially aid ENT specialists and neurosurgeons in order to approach anatomical landmarks and cranial fossas more safely during otologic and neurotologic surgeries. It could also help the design of middle ear prosthesis.     

Effect of Intensive Compared With Standard Glycemia Treatment Strategies on Mortality by Baseline Subgroup Characteristics: The Action to Control Cardiovascular Risk in Diabetes (ACCORD) trial

OBJECTIVE

To determine if baseline subgroups in the Action to Control Cardiovascular Risk in Diabetes (ACCORD) trial can be identified for whom intensive compared with standard glycemia treatment had different effects on all-cause mortality.

RESEARCH DESIGN AND METHODS

Exploratory post hoc intention-to-treat comparisons were made between intensive and standard glycemia groups on all-cause mortality by subgroups defined by baseline characteristics.

RESULTS

There were few significant interactions between baseline characteristics and effects of intensive versus standard glycemia treatment on mortality: self-reported history of neuropathy (hazard ratio [HR] 1.95, 95% CI 1.41–2.69) versus no history of neuropathy (0.99, 0.79–1.26; P value for interaction 0.0008), higher A1C (A1C >8.5%: HR 1.64, 95% CI 1.22–2.22; A1C 7.5–8.4%: 1.00, 0.75–1.34; A1C <7.5%: 1.00, 0.67–1.50; P value for interaction 0.04), and aspirin use (HR 1.45, 95% CI 1.13–1.85, compared with 0.96, 0.72–1.27, in nonusers; P value for interaction 0.03).

CONCLUSIONS

We found a remarkable similarity of effect from intensive compared with standard glycemia treatment on mortality across most baseline subgroups. No differential effect was found in subgroups defined by variables anticipated to have an interaction: age, duration of diabetes, and previous history of cardiovascular disease. The three baseline characteristics that defined subgroups for which there was a differential effect on mortality may help identify patients with type 2 diabetes at higher risk of mortality from intensive regimens for glycemic control. Further research is warranted.Numerous epidemiological studies have demonstrated a relationship between elevated A1C and a greater risk of cardiovascular (CVD) events and mortality in type 2 diabetes (1–3). Therefore, it has been hypothesized that a reduction to near-normal levels of A1C in patients with type 2 diabetes would reduce the risk of these adverse outcomes. Three large randomized controlled clinical trials testing this hypothesis in individuals with longstanding type 2 diabetes reported their main results in the past 2 years (4–6).The Data Safety Monitoring Board of the Action to Control Cardiovascular Risk in Diabetes (ACCORD) trial discontinued the intensive glycemia arm because of an increase in all-cause mortality in the intensive glycemia arm compared with the standard glycemia arm. The finding of excess mortality in the intensive arm of the ACCORD trial has led to controversy about implementation of intensive glucose control in patients with type 2 diabetes (7,8). Adding to the controversy were results of the Action in Diabetes and Vascular Disease: Preterax and Diamicron MR Controlled Evaluation (ADVANCE) and Veterans Affairs Diabetes Trial (VADT), demonstrating that although there was no significant reduction in the primary end point of CVD events, there was no increase in mortality with the intensive glycemia arm compared with the standard glycemia arm (4,6), which has raised questions about reasons for these discrepancies (9–12).A critical question relates to the applicability and generalizability of the conclusions of the ACCORD trial to the broader population or to specific subgroups of patients with type 2 diabetes. Indeed, prespecified subgroup analyses in ACCORD did suggest a significant benefit of intensive glycemic control on CVD events in those participants with lower A1C at entry or absence of CVD event by history, but there was no suggestion of a differential effect on mortality (5). However, these observations are based on only a few subgroup analyses at the time of the primary publication. The effect on mortality of intensive compared with standard glycemia treatment may have been modified by other possible characteristics of patients at entry. We have therefore carried out exploratory post hoc analyses of the effects of intensive compared with standard glycemia treatment in ACCORD participants categorized by various baseline characteristics on all-cause mortality at the time of discontinuation of the intensive glycemia treatment of ACCORD, with the goal to determine if particular subgroups at higher or lower risk from the intensive intervention can be identified.     

Adrenal Biopsy under Wide-Bore MR Imaging Guidance

Twenty-four magnetic resonance (MR) imaging–guided percutaneous adrenal biopsies performed between April 2009 and October 2016 were reviewed retrospectively. Epidemiologic, procedural, and histopathologic data were retrospectively collected. Mean size of tumors was 4.3 cm (range, 1.5–16.0 cm). Mean procedure time was 49 min (range, 24–95 min). Mean needle angulation was 27.7° (range, 0°–60°). Mean depth was 9.6 cm (range, 5.8–13.7 cm). The sensitivity, specificity, positive predictive value, negative predictive value, and accuracy of MR imaging–guided biopsy were 95.5%, 100%, 100%, 66.7%, and 95.8%, respectively. There were no immediate or delayed complications. MR imaging guidance seems safe and accurate to target adrenal-gland masses.     

Association of perturbation of oral bacterial with incident of Alzheimer's disease: A pilot study

ObjectiveThis case‐control study was designed to compare the composition of the predominant oral bacterial microbiome in Alzheimer''s disease (AD) and control group.SubjectA total of 30 adult participants (15 AD and 15 healthy individuals) were entered in this study. The composition of oral bacterial microbiome was examined by quantitative real‐time polymerase chain reaction (qPCR) using bacterial 16S rDNA gene. The levels of systemic inflammatory cytokines in both groups were assessed using enzyme‐linked immunosorbent assays (ELISA).ResultsThe loads of Porphyromonas gingivalis, Fusobacterium nucleatum, and Prevotella intermedia were significantly more abundant in the AD compared to the control group (p < 0.05). Although Aggregatibacter actinomycetemcomitans and Streptococcus mutans were relatively frequent in the AD group, no significance difference was observed in their copy number between two groups. Although the concentrations of IL‐1, IL‐6, and TNF‐α were higher in the AD group, there was a significant difference in their levels between the two groups (p < 0.05). Finally, there was a significant relationship between increased number of pathogenic bacteria in oral microbiome and higher concentration of cytokines in patient''s blood.ConclusionOur knowledge of oral microbiome and its exact association with AD is rather limited; our study showed a significant association between changes in oral microbiome bacteria, increased inflammatory cytokines, and AD.