Mathieu F. Bakhoum

TDP-43 Phosphorylation by casein kinase Iε promotes oligomerization and enhances toxicity in vivo

Dominant mutations in transactive response DNA-binding protein-43 (TDP-43) cause amyotrophic lateral sclerosis. TDP-43 inclusions occur in neurons, glia and muscle in this disease and in sporadic and inherited forms of frontotemporal lobar degeneration. Cytoplasmic localization, cleavage, aggregation and phosphorylation of TDP-43 at the Ser409/410 epitope have been associated with disease pathogenesis. TDP-43 aggregation is not a common feature of mouse models of TDP-43 proteinopathy, and TDP-43 is generally not thought to acquire an amyloid conformation or form fibrils. A number of putative TDP-43 kinases have been identified, but whether any of these functions to regulate TDP-43 phosphorylation or toxicity in vivo is not known. Here, we demonstrate that human TDP-43(Q331K) undergoes cytoplasmic localization and aggregates when misexpressed in Drosophila when compared with wild-type and M337V forms. Coexpression of Q331K with doubletime (DBT), the fly homolog of casein kinase Iε (CKIε), enhances toxicity. There is at best modest basal phosphorylation of misexpressed human TDP-43 in Drosophila, but coexpression with DBT increases Ser409/410 phosphorylation of all TDP-43 isoforms tested. Phosphorylation of TDP-43 in the fly is specific for DBT, as it is not observed using the validated tau kinases GSK-3β, PAR-1/MARK2 or CDK5. Coexpression of DBT with TDP-43(Q331K) enhances the formation of high-molecular weight oligomeric species coincident with enhanced toxicity, and treatment of recombinant oligomeric TDP-43 with rat CKI strongly enhances its toxicity in mammalian cell culture. These data identify CKIε as a potent TDP-43 kinase in vivo and implicate oligomeric species as the toxic entities in TDP-43 proteinopathies.

Ocular adnexal lymphoma: validation of American Joint Committee on Cancer seventh edition staging guidelines

Background/aims To validate the prognostic significance of the American Joint Committee on Cancer (AJCC) seventh edition staging criteria for ocular adnexal lymphoma (OAL) of all histologic subtypes. Methods Retrospective review of clinical records for all consecutive patients with OAL treated from November 1998 to December 2012. Results 130 patients were evaluated, 82 with primary and 34 with secondary OAL. Fourteen patients were excluded due to incomplete records. 71 women (61.2%) and 45 men (38.8%) had a median age of 61.5 years. Patients were followed for a median of 32.5 months. Treatment varied, in part, related to lymphoma histologic subtype. Overall, there were 17 recurrences (8 local and 9 distant) in patients with primary OAL. For primary OAL, the 5-year disease-free survival (DFS) and overall survival (OS) rates were 55.9% and 85.8%, respectively. For primary OAL, while there was a trend towards decreased 5-year DFS for more aggressive subtypes, this was not statistically significant. More advanced Ann Arbor stage was associated with decreased 5-year DFS; however, this trend was not statistically significant. However, increased AJCC seventh edition T category was associated with decreased 5-year DFS (T1=67.8%, T2=59.2%, T3=28.6%, T4=33.3%; p=0.025). Conclusion AJCC seventh edition T category was predictive of DFS in patients with OAL.

Clinical value of magnetic resonance imaging and other baseline testing for conjunctival mucosa-associated lymphoid tissue lymphoma

Abstract The objective of this study was to assess the value of magnetic resonance imaging (MRI) of the orbit for conjunctival mucosa-associated lymphoid tissue (MALT) lymphoma. The yield of other staging tests at baseline were also evaluated. Twenty-one consecutive patients treated for conjunctival MALT lymphoma were retrospectively studied. Lymphoma was staged according to both the Ann Arbor system and the seventh edition of the AJCC [American Joint Committee on Cancer] cancer staging manual. Findings on MRI of the orbit, whole-body positron emission tomography/computed tomography (PET/CT), CT of the chest/abdomen/pelvis, bone marrow (BM) biopsy and gastrointestinal (GI) endoscopy were recorded. Seventeen patients had orbital MRI. Fourteen of 17 patients (82%) with obvious conjunctival MALT lymphoma on clinical examination had a negative MRI scan. Only three patients had subtle conjunctival enhancement on orbital MRI. Ann Arbor stage at presentation was as follows: stage IE (15 patients), stage IIE (two patients) and stage IV (four patients). Eighteen of 21 patients had total-body PET/CT; four patients (22%) had hypermetabolic activity evident on PET scan. All 21 patients had bilateral BM biopsies. Fifteen of 21 patients (71%) had GI endoscopy. None of the patients had a positive BM biopsy or findings on GI endoscopy. Our data suggest that orbital MRI has a very low yield for identification of conjunctival MALT lymphoma. Clinical examination is critical in diagnosing and assessing treatment response in conjunctival MALT lymphoma. The yield for GI endoscopy and BM biopsy may also be low in staging of conjunctival MALT lymphoma.

Demise of the Flies: Why Drosophila Models Still Matter

Over the past decade, Drosophila melanogaster has emerged as a widely used model for human disease via targeted misexpression of human disease-associated proteins. The chief advantage of creating such models is that once a suitable phenotype has been obtained, the genetic toolkit of fly genetics can be used to dissect underlying disease pathways. Although some critics of this approach have argued that it has not generated many novel insights, we would argue that fly models of human neurodegenerative disorders have provided valuable information when viewed within the context of other models and systems of analysis. Here, we will provide a brief overview of some Drosophila models of neurodegenerative disorders with a special focus on our own work.

Evidence for autophagic gridlock in aging and neurodegeneration.

Autophagy is essential to neuronal homeostasis, and its impairment is implicated in the development of neurodegenerative pathology. However, the underlying mechanisms and consequences of this phenomenon remain a matter of conjecture. We show that misexpression of human tau in Drosophila induces accumulation of autophagic intermediates with a preponderance of large vacuoles, which we term giant autophagic bodies (GABs), which are reminiscent of dysfunctional autophagic entities. Lowering basal autophagy reduces GABs, whereas increasing autophagy decreases mature autolysosomes. Induction of autophagy is also associated with rescue of the tauopathy phenotype, suggesting that formation of GABs may be a compensatory mechanism rather than a trigger of neurodegeneration. Last, we show that the peculiar Biondi bodies observed in the choroid epithelium of both elderly and Alzheimers disease human brains express immunoreactive markers similar to those of GABs. Collectively, these data indicate that autophagic gridlock contributes to the development of pathology in aging and neurodegeneration.

Mitochondrial A3243G mutation results in corneal endothelial polymegathism

Purpose The mitochondrial DNA point mutation A3243G leads to a spectrum of syndromes ranging from MIDD to MELAS. Ocular manifestations include pattern macular dystrophy and concentric perifoveal atrophy. Given the high metabolic demand of corneal endothelial cells, we performed specular biomicroscopy analysis in patients harboring the mitochondrial DNA point mutation A3243G to assess for the associated presence of corneal endothelial abnormalities.

Autophagic aggregates resembling granulovacuolar degeneration in a Drosophila model of tauopathy

apoptosis following Chlamydia pneumoniae infection in neuronal cells. Dysfunctions in apoptosis and autophagy have been implicated in the neurodegeneration associated with Alzheimer’s disease (AD). Autophagy in AD pathogenesis has been shown to play a role in amyloid processing through the endosomal-lysosomal system. Apoptosis may contribute to the neuronal cell loss observed in AD; however, there is limited evidence of the apoptotic process proceeding to terminal completion.AlthoughAb1-42 has been shown to induce apoptosis in neurons and may be an early factor in AD, our previous investigations demonstrated that neurons infectedwith Chlamydia pneumoniae are resistant to apoptosis, and that Ab1-42 is induced following this infection. Thus, these studies address infection as an initiator/trigger or inhibitor for the processes of autophagy and apoptosis observed inAlzheimer’s disease. Methods: SKNMC neuronal cells obtained from ATCC were infected with the AR39 strain of Chlamydia pneumoniae at an MOI 1⁄4 1 for 24, 48, and 72 hrs and were analyzed using Real-time PCR arrays from SABiosciences specific for autophagy and apoptosis genetic markers. Results: Some major genes associated with apoptosis such as BID, DAPK1, TP53, TP73 were down regulated by 72 hrs post-infection. Genes associated with the regulation of autophagic vacuole formation such as ATG3, ATG4B, ATG4C, ATG9A, ATG9B, ATG12, IRGM, and BECN1 were upregulated within 72 hrs post-infection. With regards to genes involved with co-regulation of autophagy and apoptosis, BNIP3 was significantly up-regulated within 48-72 hrs post-infection. Of the genes linking autophagosomes to lysosomes, FAM176A was up-regulated throughout 24-72 hrs post-infection. Conclusions:Modulation of autophagy and apoptosis genes occurs in neuronal cells at 24, 48, and 72 hrs post-infection with Chlamydia pneumoniae. These genetic changes lead to dysfunction in these basic cellular processes; dysfunction in these processes has been shown to contribute to the neuropathology of late-onset Alzheimer’s disease. This work will allow future studies to further focus on the apoptotic and autophagic pathways to better understand how a pathogen such as Chlamydia pneumoniae plays a role in the development of late-onset Alzheimer’s disease.

Autologous stem cell therapy for inherited and acquired retinal disease.

The mammalian retina, derived from neural ectoderm, has little regenerative potential. For conditions where irreversible retinal pigment epithelium or photoreceptor cell loss occurs, advanced techniques are required to restore vision. Inherited retinal dystrophies and some acquired conditions, such as age-related macular degeneration, have a similar end result of photoreceptor cell death leading to debilitating vision loss. These diseases stand to benefit from future regenerative medicine as dietary recommendations and current pharmacologic therapy only seek to prevent further disease progression. Cell-based strategies, such as autologously derived induced pluripotent stem cells, have come a long way in overcoming previous technical and ethical concerns. Clinical trials for such techniques are already underway. These trials and the preceding preclinical studies will be discussed in the context of retinal disease.

Congenital grouped albinotic spots of the retinal pigment epithelium in a patient with hemihypertrophy and café au lait spots

PurposeTo describe the finding of circularly grouped hypomelanotic spots in the central macula of a patient with syndromic characteristics.MethodsCase report of a patient with albinotic spots grouped within the macula, café au lait spots, and left-sided hemihypertrophy.ResultsA 15-year-old boy presented with hypomelanotic spots which were hyperautofluorescent on fundus autofluorescence imaging with no disruption of the retinal laminae or photoreceptor inner and outer segment (IS/OS) junction on spectral domain optical coherence tomography. His developmental history included hemihypertrophy, café au lait spots over his axilla and extremities, and surgically corrected left-sided cryptorchidism. Other ocular history included resolved convergence insufficiency and red–green color blindness.ConclusionsIt is essential to recognize that circularly grouped hypomelanotic spots are a benign condition. The location and arrangement of the hypomelanotic spots were atypical for congenital grouped albinotic spots of the retinal pigment epithelium (CGAS) as they were grouped within the macula in addition to a more characteristic linear “bear track” formation in the periphery. To the authors’ knowledge, this is the first report of CGAS present in a patient with hemihypertrophy, café au lait spots, and cryptorchidism and may represent a novel syndromic association.

Paracentral Acute Middle Maculopathy and the Ischemic Cascade Associated With Retinal Vascular Occlusion

PURPOSE To analyze the spectrum of ischemia associated with paracentral acute middle maculopathy (PAMM) in eyes with acute retinal vascular occlusion and to describe an ischemic cascade. DESIGN A retrospective observational case series. METHODS Patients presenting with PAMM secondary to acute retinal vascular occlusion were identified. Analysis of multimodal imaging was performed at baseline and at follow-up visits to elucidate the patterns and progression of ischemia within the retinal layers. RESULTS Multimodal retinal imaging from 16 eyes of 16 patients with acute retinal vascular occlusion associated with PAMM was studied. Analysis of en face optical coherence tomography (OCT) segmentation of the inner nuclear layer (INL) identified distinct patterns of PAMM correlating with the severity of ischemia and not the type of occlusion. A perivenular fern-like PAMM pattern was associated with better visual outcomes (average final visual acuity was 20/25). This pattern was noted to sequentially progress in 2 cases to a diffuse globular PAMM pattern in the INL, or to a pattern of ischemia involving both the middle and inner retinal layers with commensurate vision loss. Globular patterns of PAMM or ischemia involving both the middle and inner retina correlated with poorer visual outcomes (average final visual acuity was counting fingers at 5.5 ft). These various patterns of ischemia developed in eyes with retinal vascular occlusions in which blood flow through the retinal capillary plexuses was present but was significantly reduced and delayed. CONCLUSIONS This study describes OCT findings suggestive of an ischemic cascade in eyes with retinal vascular occlusion. The middle retina at the level of the deep capillary plexus, especially at the venular pole, may be more vulnerable to ischemic injury.