Morphological and functional study of cerebral aging in rat

Aging is a physiological process characterized by a progressive decline of the “cellular homeostatic reserve”, refereed as the capability to respond suitably to exogenous and endogenous stressful stimuli. Due to their high energetic requests and post-mitotic nature, neurons are peculiarly susceptible to this phenomenon. However, the aged brain maintains a certain level of adaptive capacities and if properly stimulated may warrant a considerable functional recovery. Aim of the present research was to verify the plastic potentialities of the aging brain of rats subjected to two kind of exogenous stimuli: A) the replacement of the standard diet with a ketogenic regimen (the change forces the brain to use ketone bodies (KB) in alternative to glucose to satisfy the energetic needs) and B) a behavioural task able to induce the formation of inhibitory avoidance memory. A) Fifteen male Wistar rats of 19 months of age were divided into three groups (average body weight pair-matched), and fed for 8 weeks with different dietary regimens: i) diet containing 10% medium chain triglycerides (MCT); ii) diet containing 20% MCT; iii) standard commercial chow. Five young (5 months of age) and five old (26-27 months of age) animals fed with the standard diet were used as further controls. The following morphological parameters reflecting synaptic plasticity were evaluated in the stratum moleculare of the hippocampal CA1 region (SM CA1), in the outer molecular layer of the hippocampal dentate gyrus (OML DG), and in the granule cell layer of the cerebellar cortex (GCL-CCx): average area (S), numeric density (Nvs), and surface density (Sv) of synapses, and average volume (V), numeric density (Nvm), and volume density (Vv) of synaptic mitochondria. Moreover, succinic dehydrogenase (SDH) activity was cytochemically determined in Purkinje cells (PC) and V, Nvm, Vv, and cytochemical precipitate area/mitochondrial area (R) of SDH-positive mitochondria were evaluated. In SM CA1, MCT-KDs induced the early appearance of the morphological patterns typical of old animals: higher S and V, and lower Nvs and Nvm. On the contrary, in OML DG, Sv and Vv of MCT-KDs-fed rats were higher (as a result of higher Nvs and Nvm) vs. controls; these modifications are known to improve synaptic function and metabolic supply. The opposite effects of MCT-KDs might reflect the different susceptibility of these brain regions to the aging processes: OML DG is less vulnerable than SM CA1, and the reactivation of ketone bodies uptake and catabolism might occur more efficiently in this region, allowing the exploitation of their peculiar metabolic properties. In GCL-CCx, the results described a new scenario in comparison to that found in the hippocampal formation: 10%MCT-KD induced the early appearance of senescent patterns (decreased Nvs and Nvm; increased V), whereas 20%MCT-KD caused no changes. Since GCL-CCx is more vulnerable to age than DG, and less than CA1, these data further support the hypothesis that MCT-KDs effects in the aging brain critically depend on neuronal vulnerability to age, besides MCT percentage. Regarding PC, it was decided to evaluate only the metabolic effect of the dietetic regimen (20%MCT-KD) characterized by less side effects. KD counteracted age-related decrease in numeric density of SDH-positive mitochondria, and enhanced their energetic efficiency (R was significantly higher in MCT-KD-fed rats vs. all the controls). Since it is well known that Purkinje and dentate gyrus cells are less vulnerable to aging than CA1 neurons, these results corroborate our previous hypothesis. In conclusion, the A) experimental line provides the first evidence that morphological and functional parameters reflecting synaptic plasticity and mitochondrial metabolic competence may be modulated by MCT-KDs in the pre-senescent central nervous system, and that the effects may be heterogeneous in different brain regions. MCT-KDs seem to supply high energy metabolic intermediates and to be beneficial (“anti-aging”) for those neurons that maintain the capability to exploit them. This implies risks but also promising potentialities for the therapeutic use of these diets during aging B) Morphological parameters of synapses and synaptic mitochondria in SM CA1 were investigated in old (26-27 month-old) female Wistar rats following a single trial inhibitory avoidance task. In this memory protocol animals learn to avoid a dark compartment in which they received a mild, inescapable foot-shock. Rats were tested 3 and 6 or 9 hours after the training, divided into good and bad responders according to their performance (retention times above or below 100 s, respectively) and immediately sacrificed. Nvs, S, Sv, Nvm, V, and Vv were evaluated. In the good responder group, the numeric density of synapses and mitochondria was significantly higher and the average mitochondrial volume was significantly smaller 9 hours vs. 6 hours after the training. No significant differences were observed among bad responders. Thus, better performances in passive avoidance memory task are correlated with more efficient plastic remodeling of synaptic contacts and mitochondria in hippocampal CA1. These findings indicate that maintenance of synaptic plastic reactivity during aging is a critical requirement for preserving long-term memory consolidation.

Architectural development and dry matter production in a multisite trial on single and multiaxis apple trees (Malus domestica Borkh) grafted on different rootstocks

In two Italian sites, multiaxis trees slightly reduced primary axis length and secondary axis length of newly grafted trees, and increased the number of secondary shoots. The total length, node production, and total dry matter gain were proportional to the number of axis. Growth of both primary and secondary shoots, and dry matter accumulation, have been found to be also well related to rootstock vigour. A great variability in axillary shoot production was recorded among different environments. Grafted trees had higher primary growth, secondary axis growth, and dry matter gain than chip budded trees. Stem water potential measured in the second year after grafting was not affected by rootstocks or number of leaders. Measurements performed in New Zealand (Hawke’s Bay) during the second year after grafting revealed that both final length and growth rate of primary and secondary axis were related to the rootstock rather than to the training system. Dwarfing rootstocks reduced the number of long vegetative shoots and increased the proportion of less vigorous shoots.

A phase II, single arm study of CarbopLatin plus Etoposide with Bevacizumab and Atezolizumab in patients with exTEnded-disease small-cell lung cancer (SCLC) – CeLEBrATE trial

Small cell lung cancer (SCLC) is an aggressive neuroendocrine tumor diagnosed at extended disease SCLC (ES-SCLC) stage in about 70% of cases. The new standard of treatment for patients with ES-SCLC is a combination of platinum-etoposide chemotherapy and atezolizumab or durvalumab, two programmed cell death ligand 1 (PD-L1) inhibitory monoclonal antibodies (mAb). However, the benefit derived from the addition of PD-L1 inhibitors to chemotherapy in ES-SCLC was limited and restricted to a subset of patients. The vascular endothelial growth factor (VEGF) is the most important pro-angiogenic factor implicated in cancer angiogenesis, which is abundant in SCLC and associated with poor prognosis. Antiangiogenic agents, such as bevacizumab, a humanized mAb against VEGF, added to platinum-etoposide chemotherapy improved progression-free survival in SCLC in two trials, but it did not translate into a benefit in overall survival. Nevertheless, VEGF has also acts as a mediator of an immunosuppressive microenvironment and its inhibition can revert the immune-suppressive tumor microenvironment and potentially enhance the efficacy of immunotherapies. Based on available preclinical data, we hypothesized that VEGF inhibition by bevacizumab could improve atezolizumab efficacy in a synergistic way and designed a phase II single-arm trial of bevacizumab in combination with carboplatin, etoposide, and atezolizumab as first-line treatment in ES-SCLC. The trial, which is still ongoing, enrolled 53 patients, including those with treated or untreated asymptomatic brain metastases (provided criteria are met), who received atezolizumab, bevacizumab, carboplatin and etoposide for 4-6 cycles (induction phase), followed by maintenance with atezolizumab and bevacizumab for a maximum of 18 total cycles or until disease progression, patient refusal, unacceptable toxicity. The evaluation of efficacy of the experimental combination in terms of 1-year overall survival rate is not yet mature (primary objective of the trial). The combination was feasible and the toxicity profile manageable (secondary objective of the trial).

Aragonite-based scaffold for the treatment of knee osteochondral defects: results of a prospective multi-center trial at 2 years' follow-up

PURPOSE: To evaluate the clinical and MRI outcomes after the implantation of a nanostructured cell free aragonite-based scaffold in patients affected by knee chondral and osteochondral lesions. METHODS: 126 patients (94 men, 32 women; age 32.7±8.8 years) were included according to the following criteria: grade III or IV chondra/osteochondral lesions in the femoral condyles or throclea; 2) no limb axial deviation (i.e. varus or valgus knee > 5°); 3) no signs of knee instability; 4) no concurrent tibial or patellar chondral/osteochondral defects. All patients were treated by arthrotomic implantation of an aragonite based-scaffold by a press-fit technique. Patients were prospectively evaluated by IKDC, Tegner, Lysholm and KOOS scores preoperatively and then at 6, 12, 18 and 24-months follow-up. MRI was also performed to evaluate the amount of defect filling by regenerated cartilage. Failures were defined as the need for re-intervention in the index knee within the follow-up period. RESULTS: Average defect size was 2±1.3 cm2 and in most cases a single scaffold was used. A significant improvement in each clinical score was recorded from basal level to 24 months’ follow-up. In particular, the IKDC subjective score increased from 42.14±16 to 70.94±24.69 and the Tegner score improved from 2.95±1.90 to 4.82±1.85 (p<0.0005). Lysholm score and all the subscales of KOOS showed a similar trend over time. Age of the patient at implantation, size of the defect and BMI were correlated with lower clinical outcome. The presence of OA didn’t influence the clinical results. MRI evaluation showed a significant increase in defect filling over time, with the highest value reached at 24 months. Failures occurred in eleven patients (8.7%). CONCLUSION: The aragonite-based biomimetic osteochondral scaffold proved to be safe, and encouraging clinical and radiographic outcomes were documented up to 2 years’ follow-up.