Intravital Imaging of a Massive Lymphocyte Response in the Cortical Dura of Mice after Peripheral Infection by Trypanosomes

Peripheral infection by Trypanosoma brucei, the protozoan responsible for sleeping sickness, activates lymphocytes, and, at later stages, causes meningoencephalitis. We have videoed the cortical meninges and superficial parenchyma of C56BL/6 reporter mice infected with T.b.brucei. By use of a two-photon microscope to image through the thinned skull, the integrity of the tissues was maintained. We observed a 47-fold increase in CD2+ T cells in the meninges by 12 days post infection (dpi). CD11c+ dendritic cells also increased, and extravascular trypanosomes, made visible either by expression of a fluorescent protein, or by intravenous injection of furamidine, appeared. The likelihood that invasion will spread from the meninges to the parenchyma will depend strongly on whether the trypanosomes are below the arachnoid membrane, or above it, in the dura. Making use of optical signals from the skull bone, blood vessels and dural cells, we conclude that up to 40 dpi, the extravascular trypanosomes were essentially confined to the dura, as were the great majority of the T cells. Inhibition of T cell activation by intraperitoneal injection of abatacept reduced the numbers of meningeal T cells at 12 dpi and their mean speed fell from 11.64 ± 0.34 μm/min (mean ± SEM) to 5.2 ± 1.2 μm/min (p = 0.007). The T cells occasionally made contact lasting tens of minutes with dendritic cells, indicative of antigen presentation. The population and motility of the trypanosomes tended to decline after about 30 dpi. We suggest that the lymphocyte infiltration of the meninges may later contribute to encephalitis, but have no evidence that the dural trypanosomes invade the parenchyma.     

Stereotaxic Infusion of Oligomeric Amyloid-beta into the Mouse Hippocampus

Alzheimer’s disease is a neurodegenerative disease affecting the aging population. A key neuropathological feature of the disease is the over-production of amyloid-beta and the deposition of amyloid-beta plaques in brain regions of the afflicted individuals. Throughout the years scientists have generated numerous Alzheimer’s disease mouse models that attempt to replicate the amyloid-beta pathology. Unfortunately, the mouse models only selectively mimic the disease features. Neuronal death, a prominent effect in the brains of Alzheimer’s disease patients, is noticeably lacking in these mice. Hence, we and others have employed a method of directly infusing soluble oligomeric species of amyloid-beta - forms of amyloid-beta that have been proven to be most toxic to neurons - stereotaxically into the brain. In this report we utilize male C57BL/6J mice to document this surgical technique of increasing amyloid-beta levels in a select brain region. The infusion target is the dentate gyrus of the hippocampus because this brain structure, along with the basal forebrain that is connected by the cholinergic circuit, represents one of the areas of degeneration in the disease. The results of elevating amyloid-beta in the dentate gyrus via stereotaxic infusion reveal increases in neuron loss in the dentate gyrus within 1 week, while there is a concomitant increase in cell death and cholinergic neuron loss in the vertical limb of the diagonal band of Broca of the basal forebrain. These effects are observed up to 2 weeks. Our data suggests that the current amyloid-beta infusion model provides an alternative mouse model to address region specific neuron death in a short-term basis. The advantage of this model is that amyloid-beta can be elevated in a spatial and temporal manner.     

Complex offspring size effects: variations across life stages and between species

Classical optimality models of offspring size and number assume a monotonically increasing relationship between offspring size and performance. In aquatic organisms with complex life cycles, the size–performance function is particularly hard to grasp because measures of performance are varied and their relationships with size may not be consistent throughout early ontogeny. Here, we examine size effects in premetamorphic (larval) and postmetamorphic (juvenile) stages of brooding marine animals and show that they vary contextually in strength and direction during ontogeny and among species. Larger offspring of the sea anemone Urticina felina generally outperformed small siblings at the larval stage (i.e., greater settlement and survival rates under suboptimal conditions). However, results differed when analyses were conducted at the intrabrood versus across‐brood levels, suggesting that the relationship between larval size and performance is mediated by parentage. At the juvenile stage (15 months), small offspring were less susceptible than large ones to predation by subadult nudibranchs and both sizes performed similarly when facing adult nudibranchs. In a sympatric species with a different life history (Aulactinia stella), all juveniles suffered similar predation rates by subadult nudibranchs, but smaller juveniles performed better (lower mortalities) when facing adult nudibranchs. Size differences in premetamorphic performance of U. felina were linked to total lipid contents of larvae, whereas size‐specific predation of juvenile stages followed the general predictions of the optimal foraging strategy. These findings emphasize the challenge in gathering empirical support for a positive monotonic size–performance function in taxa that exhibit complex life cycles, which are dominant in the sea.     

Diet Is Critical for Prolonged Glycemic Control after Short-Term Insulin Treatment in High-Fat Diet-Induced Type 2 Diabetic Male Mice

Background

Clinical studies suggest that short-term insulin treatment in new-onset type 2 diabetes (T2DM) can promote prolonged glycemic control. The purpose of this study was to establish an animal model to examine such a “legacy” effect of early insulin therapy (EIT) in long-term glycemic control in new-onset T2DM. The objective of the study was to investigate the role of diet following onset of diabetes in the favorable outcomes of EIT.

Methodology

As such, C57BL6/J male mice were fed a high-fat diet (HFD) for 21 weeks to induce diabetes and then received 4 weeks of daily insulin glargine or sham subcutaneous injections. Subsequently, mice were either kept on the HFD or switched to a low-fat diet (LFD) for 4 additional weeks.

Principal Findings

Mice fed a HFD gained significant fat mass and displayed increased leptin levels, increasing insulin resistance (poor HOMA-IR) and worse glucose tolerance test (GTT) performance in comparison to mice fed a LFD, as expected. Insulin-treated diabetic mice but maintained on the HFD demonstrated even greater weight gain and insulin resistance compared to sham-treated mice. However, insulin-treated mice switched to the LFD exhibited a better HOMA-IR compared to those mice left on a HFD. Further, between the insulin-treated and sham control mice, in spite of similar HOMA-IR values, the insulin-treated mice switched to a LFD following insulin therapy did demonstrate significantly better HOMA-B% values than sham control and insulin-treated HFD mice.

Conclusion/Interpretation

Early insulin treatment in HFD-induced T2DM in C57BL6/J mice was only beneficial in animals that were switched to a LFD after insulin treatment which may explain why a similar legacy effect in humans is achieved clinically in only a portion of cases studied, emphasizing a vital role for diet adherence in diabetes control.     

Relationship between Fungal Colonisation of the Respiratory Tract in Lung Transplant Recipients and Fungal Contamination of the Hospital Environment

Background

Aspergillus colonisation is frequently reported after lung transplantation. The question of whether aspergillus colonisation is related to the hospital environment is crucial to prevention.

Method

To elucidate this question, a prospective study of aspergillus colonisation after lung transplantation, along with a mycological survey of the patient environment, was performed.

Results

Forty-four consecutive patients were included from the day of lung transplantation and then examined weekly for aspergillus colonisation until hospital discharge. Environmental fungal contamination of each patient was followed weekly via air and surface sampling. Twelve patients (27%) had transient aspergillus colonisation, occurring 1–13 weeks after lung transplantation, without associated manifestation of aspergillosis. Responsible Aspergillus species were A. fumigatus (6), A. niger (3), A. sydowii (1), A. calidoustus (1) and Aspergillus sp. (1). In the environment, contamination by Penicillium and Aspergillus was predominant. Multivariate analysis showed a significant association between occurrence of aspergillus colonisation and fungal contamination of the patient’s room, either by Aspergillus spp. in the air or by A.fumigatus on the floor. Related clinical and environmental isolates were genotyped in 9 cases of aspergillus colonisation. For A. fumigatus (4 cases), two identical microsatellite profiles were found between clinical and environmental isolates collected on distant dates or locations. For other Aspergillus species, isolates were different in 2 cases; in 3 cases of aspergillus colonisation by A. sydowii, A. niger and A. calidoustus, similarity between clinical and environmental internal transcribed spacer and tubulin sequences was >99%.

Conclusion

Taken together, these results support the hypothesis of environmental risk of hospital acquisition of aspergillus colonisation in lung transplant recipients.     

The Amelioration of Hepatic Steatosis by Thyroid Hormone Receptor Agonists Is Insufficient to Restore Insulin Sensitivity in Ob/Ob Mice

Thyroid hormone receptor (TR) agonists have been proposed as therapeutic agents to treat non-alcoholic fatty liver disease (NAFLD) and insulin resistance. We investigated the ability of the TR agonists GC-1 and KB2115 to reduce hepatic steatosis in ob/ob mice. Both compounds markedly reduced hepatic triglyceride levels and ameliorated hepatic steatosis. However, the amelioration of fatty liver was not sufficient to improve insulin sensitivity in these mice and reductions in hepatic triglycerides did not correlate with improvements in insulin sensitivity or glycemic control. Instead, the effects of TR activation on glycemia varied widely and were found to depend upon the time of treatment as well as the compound and dosage used. Lower doses of GC-1 were found to further impair glycemic control, while a higher dose of the same compound resulted in substantially improved glucose tolerance and insulin sensitivity, despite all doses being equally effective at reducing hepatic triglyceride levels. Improvements in glycemic control and insulin sensitivity were observed only in treatments that also increased body temperature, suggesting that the induction of thermogenesis may play a role in mediating these beneficial effects. These data illustrate that the relationship between TR activation and insulin sensitivity is complex and suggests that although TR agonists may have value in treating NAFLD, their effect on insulin sensitivity must also be considered.     

What Do Eye Gaze Metrics Tell Us about Motor Imagery?

Many of the brain structures involved in performing real movements also have increased activity during imagined movements or during motor observation, and this could be the neural substrate underlying the effects of motor imagery in motor learning or motor rehabilitation. In the absence of any objective physiological method of measurement, it is currently impossible to be sure that the patient is indeed performing the task as instructed. Eye gaze recording during a motor imagery task could be a possible way to “spy” on the activity an individual is really engaged in. The aim of the present study was to compare the pattern of eye movement metrics during motor observation, visual and kinesthetic motor imagery (VI, KI), target fixation, and mental calculation. Twenty-two healthy subjects (16 females and 6 males), were required to perform tests in five conditions using imagery in the Box and Block Test tasks following the procedure described by Liepert et al. Eye movements were analysed by a non-invasive oculometric measure (SMI RED250 system). Two parameters describing gaze pattern were calculated: the index of ocular mobility (saccade duration over saccade + fixation duration) and the number of midline crossings (i.e. the number of times the subjects gaze crossed the midline of the screen when performing the different tasks). Both parameters were significantly different between visual imagery and kinesthesic imagery, visual imagery and mental calculation, and visual imagery and target fixation. For the first time we were able to show that eye movement patterns are different during VI and KI tasks. Our results suggest gaze metric parameters could be used as an objective unobtrusive approach to assess engagement in a motor imagery task. Further studies should define how oculomotor parameters could be used as an indicator of the rehabilitation task a patient is engaged in.     

Impact of Placental Plasmodium falciparum Malaria on the Profile of Some Oxidative Stress Biomarkers in Women Living in Yaoundé, Cameroon

Background

Impact of the pathophysiology of Plasmodium falciparum placental malaria (PM) on the profile of some oxidative stress biomarkers and their relationship with poor pregnancy outcomes in women remain unknown.

Methods

Between 2013 and 2014, peripheral blood and placenta tissue from 120 Cameroonian women at delivery were assessed for maternal haemoglobin and, parasitaemia respectively. Parasite accumulation in the placenta was investigated histologically. The levels of oxidative stress biomarkers Malondialdehyde (MDA), Nitric Oxide (NO), Superoxide dismutase (SOD), Catalase (CAT) and Gluthatione (GSH) in the supernatant of teased placenta tissues were determined by Colorimetric enzymatic assays.

Results

Parasitaemia was inversely related to haemoglobin levels and birth weight (P <0.001 and 0.012, respectively). The level of lipid peroxide product (MDA) was significantly higher in the malaria infected (P = 0.0047) and anaemic (P = 0.024) women compared to their non-infected and non-anaemic counterparts, respectively. A similar trend was observed with SOD levels, though not significant. The levels of MDA also correlated positively with parasitaemia (P = 0.0024) but negatively with haemoglobin levels (P = 0.002). There was no association between parasitaemia, haemoglobin level and the other oxidative stress biomarkers. From histological studies, levels of MDA associated positively and significantly with placenta malaria infection and the presence of malaria pigments. The levels of SOD, NO and CAT increased with decreasing leukocyte accumulation in the intervillous space. Baby birth weight increased significantly with SOD and CAT levels, but decreased with levels of GSH.

Conclusions

Placental P. falciparum infection may cause oxidative stress of the placenta tissue with MDA as a potential biomarker of PM, which alongside GSH could lead to poor pregnancy outcomes (anaemia and low birth weight). This finding contributes to the understanding of the pathophysiology of P. falciparum placental malaria in women.     

Knockout Serum Replacement Promotes Cell Survival by Preventing BIM from Inducing Mitochondrial Cytochrome C Release

Knockout serum replacement (KOSR) is a nutrient supplement commonly used to replace serum for culturing stem cells. We show here that KOSR has pro-survival activity in chronic myelogenous leukemia (CML) cells transformed by the BCR-ABL oncogene. Inhibitors of BCR-ABL tyrosine kinase kill CML cells by stimulating pro-apoptotic BIM and inhibiting anti-apoptotic BCL2, BCLxL and MCL1. We found that KOSR protects CML cells from killing by BCR-ABL inhibitors—imatinib, dasatinib and nilotinib. The protective effect of KOSR is reversible and not due to the selective outgrowth of drug-resistant clones. In KOSR-protected CML cells, imatinib still inhibited the BCR-ABL tyrosine kinase, reduced the phosphorylation of STAT, ERK and AKT, down-regulated BCL2, BCLxL, MCL1 and up-regulated BIM. However, these pro-apoptotic alterations failed to cause cytochrome c release from the mitochondria. With mitochondria isolated from KOSR-cultured CML cells, we showed that addition of recombinant BIM protein also failed to cause cytochrome c release. Besides the kinase inhibitors, KOSR could protect cells from menadione, an inducer of oxidative stress, but it did not protect cells from DNA damaging agents. Switching from serum to KOSR caused a transient increase in reactive oxygen species and AKT phosphorylation in CML cells that were protected by KOSR but not in those that were not protected by this nutrient supplement. Treatment of KOSR-cultured cells with the PH-domain inhibitor MK2206 blocked AKT phosphorylation, abrogated the formation of BIM-resistant mitochondria and stimulated cell death. These results show that KOSR has cell-context dependent pro-survival activity that is linked to AKT activation and the inhibition of BIM-induced cytochrome c release from the mitochondria.