Jörg C. Schmidt

Activation of TLR2 and TLR4 by Glycosylphosphatidylinositols Derived from Toxoplasma gondii

GPIs isolated from Toxoplasma gondii, as well as a chemically synthesized GPI lacking the lipid moiety, activated a reporter gene in Chinese hamster ovary cells expressing TLR4, while the core glycan and lipid moieties cleaved from the GPIs activated both TLR4- and TLR2-expressing cells. MyD88, but not TLR2, TLR4, or CD14, is absolutely needed to trigger TNF-α production by macrophages exposed to T. gondii GPIs. Importantly, TNF-α response to GPIs was completely abrogated in macrophages from TLR2/4-double-deficient mice. MyD88−/− mice were more susceptible to death than wild-type (WT), TLR2−/−, TLR4−/−, TLR2/4−/−, and CD14−/− mice infected with the ME-49 strain of T. gondii. The cyst number was higher in the brain of TLR2/4−/−, but not TLR2−/−, TLR4−/−, and CD14−/−, mice, as compared with WT mice. Upon infection with the ME-49 strain of T. gondii, we observed no decrease of IL-12 and IFN-γ production in TLR2-, TLR4-, or CD14-deficient mice. Indeed, splenocytes from T. gondii-infected TLR2−/− and TLR2/4−/− mice produced more IFN-γ than cells from WT mice in response to in vitro stimulation with parasite extracts enriched in GPI-linked surface proteins. Together, our results suggest that both TLR2 and TLR4 receptors may participate in the host defense against T. gondii infection through their activation by the GPIs and could work together with other MyD88-dependent receptors, like other TLRs or even IL-18R or IL-1R, to obtain an effective host response against T. gondii infection.

Management of acute submacular hemorrhage using recombinant tissue plasminogen activator and gas

Abstract · Purpose: To assess the effects of intravitreal injection of recombinant tissue plasminogen activator (rTPA) and gas on submacular hemorrhage in age-related macular degeneration (ARMD). · Methods: Eleven consecutive patients (11 eyes) with subretinal hemorrhage due to ARMD involving the fovea with elevation of the neurosensory retina were included in this study. Subretinal hemorrhage occured 12 h to 14 days before onset of therapy. Injection of rTPA through the pars plana in a dose of 50 or 100 μg was performed. Gas instillation (0.2–0.4 ml) followed rTPA injection, either immediately after injection (7 patients) or during the following day (4 patients). · Results: After intravitreal injection of rTPA, subretinal clots were totally or partially liquefied when treatment started up to 3 days after onset of bleeding. In all patients treated with 100 μg rTPA a large exudative retinal detachment of the inferior retina resulted, which reabsorbed spontaneously within 2 weeks. After reattachment of the exudative retinal detachment hyperpigmentation of the retinal pigment epithelium was noted. Temporary opacification of the vitreous was observed between the 2nd and 7th postoperative day in 5 eyes (45.5%). Postoperative visual acuity increased in 5 patients (45.5%). · Conclusion: Intravitreal application of rTPA followed by gas injection is a sufficient and convenient technique for effective removal of freshly formed submacular hemorrhage. Removal is mediated through combined enzymatic (rTPA) and mechanical (gas) effects. This technique offers a quick recovery of vision in eyes with less severe ARMD.

Acute retinal pigment epithelial tear following intravitreal bevacizumab (Avastin) injection for occult choroidal neovascularisation secondary to age related macular degeneration

Retinal pigment epithelium (RPE) tears are well recognised complications of pigment epithelial detachments (PED) in age related macular degeneration (AMD) and may arise spontaneously after trauma, photocoagulation, or photodynamic therapy (PDT).1 Rosenfeld et al recently reported favourable results after intravitreal (IV) bevacizumab (Avastin) injection in neovascular AMD.2 We present two patients, who developed an RPE tear after an intravitreal Avastin injection. The first case was a 64 year old man with an occult CNV with a PED in the right eye ( fig 1A–C). His visual acuity (VA) gradually declined from 20/30 to 20/60. Four days after an uneventful IV injection of 0.05 ml Avastin, the patient noted a sudden drop in VA. His VA was 20/80 while fluorescein angiography (FA) and optical coherence tomography revealed a large RPE tear (fig 2A–C). The second case was a 84 year old woman with an occult CNV with a PED. Her VA was 20/60 when she required an IV Avastin injection. When …

Spontaneous separation of epiretinal membrane in young subjects: personal observations and review of the literature

BackgroundIdiopathic epiretinal membranes (ERM) grow on the surface of the internal limiting membrane (ILM) and are a very uncommon condition in young subjects.MethodsWe report six young subjects with ERM and describe the spontaneous separation of the membranes. The functional and anatomical recovery was assessed by Snellen visual acuity, Amsler grid and funduscopy. Selected cases were additionally assessed by optical coherence tomography (OCT).ResultsAll subjects initially claimed a sudden, unilateral, reduction in visual acuity (VA) with severe distortion at the age of 15–30 years. No ocular trauma or disease was ascertained in any case. Initial fundus examination demonstrated a gray–whitish ERM with translucent stress lines over the macula. Visual recovery occurred in all cases after resolution of the ERM. Sequential OCT demonstrated the successive peeling of an ERM accompanied by normalization of foveal thickness.ConclusionsYoung subjects should be counseled about the favorable prognosis for maintaining good vision and possible spontaneous membrane separation. Conservative observation is advocated if the visual disturbance is located temporally, as functional recovery and spontaneous membrane separation may occur. When the contracting forces of the immature ERM are stronger than its adhesions to the retina, the membrane may separate spontaneously.

Incidence of rhegmatogenous retinal detachments after intravitreal antivascular endothelial factor injections

Acta Ophthalmol. 2011: 89: 70–75

Patent Blue: A Novel Vital Dye in Vitreoretinal Surgery

Purpose: To determine a novel vital dye (patent blue; Blueron®) for vitreoretinal surgery in a prospective consecutive case series. Methods: Five patients with either idiopathic epiretinal membrane (ERM; n = 2), proliferative vitreoretinopathy (n = 2), or a macular hole (n = 1) underwent a three-port pars plana vitrectomy. Patent blue assisted staining of the retinal surface followed by a consecutive peeling of the ERM (n = 4) or of the internal limiting membrane (ILM; n = 1) was performed. The main outcome measures were quality of intraoperative visualization of preretinal structures and postoperative visual acuity. Results: The dye induced a moderate staining (++) of the ERM and a mild staining (+) of the ILM. Complete ERM and ILM removal was successfully achieved in all cases. A mean visual improvement of three Snellen lines was observed 6 months postoperatively. No visual field defects or visible retinal pigment epithelial changes were present 6 months postoperatively. Conclusion: Patent blue, a novel dye for intraocular applications, may be added as an alternative dye in chromovitrectomy.

A Highly Immunogenic and Protective Middle East Respiratory Syndrome Coronavirus Vaccine Based on a Recombinant Measles Virus Vaccine Platform

ABSTRACT In 2012, the first cases of infection with the Middle East respiratory syndrome coronavirus (MERS-CoV) were identified. Since then, more than 1,000 cases of MERS-CoV infection have been confirmed; infection is typically associated with considerable morbidity and, in approximately 30% of cases, mortality. Currently, there is no protective vaccine available. Replication-competent recombinant measles virus (MV) expressing foreign antigens constitutes a promising tool to induce protective immunity against corresponding pathogens. Therefore, we generated MVs expressing the spike glycoprotein of MERS-CoV in its full-length (MERS-S) or a truncated, soluble variant of MERS-S (MERS-solS). The genes encoding MERS-S and MERS-solS were cloned into the vaccine strain MVvac2 genome, and the respective viruses were rescued (MVvac2-CoV-S and MVvac2-CoV-solS). These recombinant MVs were amplified and characterized at passages 3 and 10. The replication of MVvac2-CoV-S in Vero cells turned out to be comparable to that of the control virus MVvac2-GFP (encoding green fluorescent protein), while titers of MVvac2-CoV-solS were impaired approximately 3-fold. The genomic stability and expression of the inserted antigens were confirmed via sequencing of viral cDNA and immunoblot analysis. In vivo, immunization of type I interferon receptor-deficient (IFNAR−/−)-CD46Ge mice with 2 × 105 50% tissue culture infective doses of MVvac2-CoV-S(H) or MVvac2-CoV-solS(H) in a prime-boost regimen induced robust levels of both MV- and MERS-CoV-neutralizing antibodies. Additionally, induction of specific T cells was demonstrated by T cell proliferation, antigen-specific T cell cytotoxicity, and gamma interferon secretion after stimulation of splenocytes with MERS-CoV-S presented by murine dendritic cells. MERS-CoV challenge experiments indicated the protective capacity of these immune responses in vaccinated mice. IMPORTANCE Although MERS-CoV has not yet acquired extensive distribution, being mainly confined to the Arabic and Korean peninsulas, it could adapt to spread more readily among humans and thereby become pandemic. Therefore, the development of a vaccine is mandatory. The integration of antigen-coding genes into recombinant MV resulting in coexpression of MV and foreign antigens can efficiently be achieved. Thus, in combination with the excellent safety profile of the MV vaccine, recombinant MV seems to constitute an ideal vaccine platform. The present study shows that a recombinant MV expressing MERS-S is genetically stable and induces strong humoral and cellular immunity against MERS-CoV in vaccinated mice. Subsequent challenge experiments indicated protection of vaccinated animals, illustrating the potential of MV as a vaccine platform with the potential to target emerging infections, such as MERS-CoV.

Incidence of Damage to the Crystalline Lens During Intravitreal Injections

PURPOSE To report the incidence of traumatic lens injuries as a complication of intravitreal injection at 5 high-volume academic centers. METHODS We determined in a retrospective, interventional, multicenter case series the consecutive number of the injections between January 5, 2006 and December 22, 2008 from the injection log books. All injections were performed under sterile conditions in a laying position, 3.5-4.0 mm behind the limbus in an oblique fashion. The main outcome measure was the incidence of lens damage. RESULTS A total of 32,318 intravitreal injections were performed, and 3 cases of iatrogenic lens damage were reported during 36 consecutive months. All affected eyes were hyperopic. The overall incidence rate of lens injury was 0.006% (2/32,318) for intravitreal injections and 1 during a paracentesis 0.003 (1/32,318). The rate of phakic eyes determined was 67%, and thus, the incidence rate of lens damage in phakic eyes was 0.009% (2/21,653) (95% confidence interval, 0.00%-0.05%). CONCLUSIONS Although there is no agreement regarding the proper intravitreal injection technique, the incidence of traumatic injuries to the crystalline lens was very low in a large series of injected patients in a community setting. The incidence compares favorably with that reported in clinical trials in which much more extensive preinjection preparation was mandated. A good preparation of the surgical incision with proper anesthesia and detailed information of the patient, as well as good anatomical skills of the treating physician, are mandatory to prevent this rare adverse event.

High incidence of vitreomacular traction in recurrent choroidal neovascularisation after repeated photodynamic therapy

Background: The causes of recurrent choroidal neovascularisation (CNV) after photodynamic therapy (PDT) remain controversial. Subretinal surgery was carried out after unsuccessful PDT. Aims: To determine intraoperatively the status of the posterior vitreous interface. Design: Interventional case series. Methods: Conventional three-port vitrectomy was carried out in 10 eyes with CNV that had undergone 1–4 PDT sessions. The vitreous cutter was held close to the edge of the optic nerve to evaluate the status of the posterior vitreous. Results: Lesion size showed an increase from 1.5 (standard deviation (SD) 0.53) to 2.3 (SD 0.83) macular photocoagulation study disc diameters, between the first and the last PDT. Intraoperative findings during vitrectomy showed little liquefaction of the vitreous gel and an incomplete posterior-vitreous detachment, with remarkably firm attachments at the macula in all cases (10/10). Conclusion: We determined an abnormally high incidence of vitreous attachments in eyes with recurrent CNV. Vitreomacular attachments may trigger the progression or recurrence of CNV.

Fatty Acids from Plasmodium falciparum Down-Regulate the Toxic Activity of Malaria Glycosylphosphatidylinositols

ABSTRACT Plasmodium falciparum malaria kills roughly 2.5 million people, mainly children, annually. Much of this mortality is thought to arise from the actions of a malarial toxin. This toxin, identified as glycosylphosphatidylinositol (GPI), is a major pathogenicity determinant in malaria. A malarial molecule, Pfj, labeled by [3H]glucosamine like the GPIs, was identified as a non-GPI molecule. Here we show that Pfj is able to down-regulate tumor necrosis factor alpha (TNF-α) production induced by the GPI of P. falciparum. Mass spectrometry analysis showed that Pfj was not a single molecule but represented a number of molecules. Separation methods, such as cation-exchange chromatography and thin-layer chromatography, were used to isolate and identify the following four main fatty acids responsible for the inhibitory effect on TNF-α production: myristic, pentadecanoic, palmitic, and palmitoleic acids. This regulatory effect on cytokine production suggests that there is balanced bioactivity for the different categories of malarial lipids.