Biomedical subjects

Magdalene J Seiler

Publications and source records attributed to Magdalene J Seiler.

17 recordsLinked to original sources

PubMed2026

Generation of a New Immunodeficient Rat Model of Retinal Degeneration With LSL TdTomato Reporter and TdTomato-Pcp2 Expression.

PURPOSE: The purpose of this study was to develop a fluorescently labeled immunodeficient retinal degenerate (RD) rat model for studying photoreceptor degeneration and transplant-host connectivity using the Cre-lox system. METHODS: We developed gene constructs for CAG-LSL-TdTomato (expressing floxed TdTomato) and Pcp2-Cre (marker for ON-bipolar cells) that were injected into rat embryos. The LSL TdTomato reporter strain, created on immunodeficient RhoS334ter-3 rats (RRRC #539), was bred to homozygosity (strain SD-Foxn1rnuTg((Rho-S334X)3,CAG-TdTomato)1010Mjsuc, RRRC #1055, "RNT"). The gene construct Pcp2-Cre was injected into Long-Evans (LE) rat embryos, resulting in two Pcp2-cre founders (strain PCP2 Cre-1105 RKI, "Pcp2"), with targeted and targeted/random insertion of the transgene. F1 offspring were bred to homozygosity and immunodeficiency. To test whether TdTomato expression can be induced in "RNT" rats expressing floxed TdTomato, retinal explants of P9 "RNT" rats were exposed to AAV-PHP.eB-hSyn-myc-Cre (AAV-Syn-Cre) virus. Homozygous rats of both strains ("RNT" and Pcp2-Cre) were crossbred to generate RD TdTomato-Pcp2 ("RTP") rats. Retinas were stained for various retinal markers. GFP-expressing rat retinas were transplanted to 6-week-old "RTP" rats and analyzed after 37 and 77 days. RESULTS: AAV-Syn-Cre induced TdTomato expression in "RNT" retinas. TdTomato-Pcp2 RD rats developed RD similar to the original Rho S334ter-3 rats. Retinas with targeted Pcp2-Cre insertion showed TdTomato in retinal interneurons, overlapping with Pcp2-staining ON bipolar cells, and cones. Retinas with random Pcp2-Cre insertion exhibited additional TdTomato in many other cells. Pcp2-TdTomato expression defined transplant-host boundaries. CONCLUSIONS: We created a unique RD rat model for studying retinal transplant connectivity which can also be used to generate RD rats with other cell-specific labels. TRANSLATIONAL RELEVANCE: This newly created rat is useful for cell therapy and retinal degeneration studies.

Animals↗

PubMed2005

Light response differences in the superior colliculus of albino and pigmented rats.

Multi-unit visual responses to light intensities ranging from -6.46 to 0.81 logcd/m2 were recorded from the surface of the superior colliculus of dark-adapted normal pigmented and normal albino rats. Light sensitivity was significantly higher in albinos. The response onset latency was inversely proportional to the stimulus intensity. The progression of the stimulus intensity versus response onset latency curve showed a considerable difference between pigmented and albino rats. At low light levels, longer response onset latencies were recorded in pigmented rats than in albinos. This can be attributed to the transmission of rod-driven responses. The differences observed in the light response characteristics of albino rats may be indicative of their visual abnormalities.

Action Potentials↗

PubMed2005

Retinal transplants evaluated by optical coherence tomography in photoreceptor degenerate rats.

Optical coherence tomography (OCT), a non-invasive method, was used for qualitative assessment of fetal retinal sheet transplants by non-invasive imaging. Rhodopsin-mutant S334ter-line-3 rats with fast retinal degeneration (28-37-day old) were transplanted with fetal retinal sheets from embryonic day (E) 18-19 pigmented normal rats. Retinal thickness measurements from transplanted (n = 51), no surgery control (n = 8), and normal pigmented rat eyes (n = 6) were obtained using a Zeiss stratus OCT-3 scanning instrument. Frozen retinal sections were stained with hematoxylin/eosin. S334ter-line-3 rats showed significant reduction in OCT retinal thickness (p<0.001) compared to normal pigmented rats at the age of 21 days. In 62% of the transplanted rats, OCT scanning revealed the presence of a subretinal graft, which was confirmed by subsequent histology. Retinal thickness in the transplant area was significantly increased compared to the area outside the transplant and to non-transplanted eyes (p<0.001). While most of the transplants with single-band OCT images (87%) had rosetted transplants, a considerable proportion of transplants having a multi-band OCT image were found to have well-laminated areas in the graft after histological evaluation. Following retinal transplantation in rodents, OCT imaging data correlated mostly with transplant morphology. OCT is a useful technique for in vivo screening and evaluation of retinal transplants. This technique determines surgical outcomes at a much earlier stage.

Animals↗

PubMed2005

Photoreceptor differentiation and integration of retinal progenitor cells transplanted into transgenic rats.

Previous studies evaluating neural stem cells transplanted into the mature retina have demonstrated limited levels of graft-host integration and photoreceptor differentiation. The purpose of this investigation is to enhance photoreceptor cell differentiation and integration of retinal progenitor cells (RPC) following subretinal transplantation into retinal degenerate rats by optimization of isolation, expansion, and transplantation procedures. RPCs were isolated from human placental alkaline phosphatase (hPAP)-positive embryonic day 17 (E17) rat retina and expanded in serum-free defined media. RPCs at passage 2 underwent in vitro induction with all trans retinoic acid or were transplanted into the subretinal space of post-natal day (P) 17 S334ter-3 and S334ter-5 transgenic rats. Animals were examined post-operatively by ophthalmoscopy and optical coherence tomography (OCT) at weeks 1 and 4. Differentiation profiles of RPCs, both in vitro and in vivo were analysed microscopically by immunohistochemistry for various retinal cell specific markers. Our results demonstrated that the majority of passage 2 RPCs differentiated into retina-specific neurons expressing rhodopsin after in vitro induction. Following subretinal transplantation, grafted cells formed a multi-layer cellular sheet in the subretinal space in both S334ter-3 and S334ter-5 rats. Prominent retina-specific neuronal differentiation was observed in both rat lines as evidenced by recoverin or rhodopsin staining in 80% of grafted cells. Less than 5% of the grafted cells expressed glial fibrillary acidic protein. Synapsin-1 (label for nerve terminals) positive neural processes were present at the graft-host interface. Expression profiles of the grafted RPCs were similar to those of RPCs induced to differentiate in vitro using all-trans retinoic acid. In contrast to our previous study, grafted RPCs can demonstrate extensive rhodopsin expression, organize into layers, and show some features of apparent integration with the host retina following subretinal transplantation in slow and fast retinal degenerate rats. The similarity of the in vitro and in vivo RPC differentiation profiles suggests that intrinsic signals may have a significant contribution to RPC cell fate determination.

Animals↗

PubMed2005

Transsynaptic virus tracing from host brain to subretinal transplants.

The aim of this study was to establish synapses between a transplant and a degenerated retina. To tackle this difficult task, a little-known but well-established CNS method was chosen: trans-synaptic pseudorabies virus (PRV) tracing. Sheets of E19 rat retina with or without retinal pigment epithelium (RPE) were transplanted to the subretinal space in 33 Royal College of Surgeons (RCS) and transgenic s334ter-5 rats with retinal degeneration. Several months later, PRV-BaBlu (expressing E. colibeta-galactosidase) or PRV-Bartha was injected into an area of the exposed superior colliculus (SC), topographically corresponding to the transplant placement in the retina. Twenty normal rats served as controls. After survival times of 1-5 days, retinas were examined for virus by X-gal histochemistry, immunohistochemistry and electron microscopy. In normal controls, virus was first seen in retinal ganglion cells and Müller glia after 1-1.5 days, and had spread to all retinal layers after 2-3 days. Virus-labeled cells were found in 16 of 19 transplants where the virus injection had retrogradely labeled the topographically correct transplant area of the host retina. Electron microscopically, enveloped and nonenveloped virus could clearly be detected in infected cells. Enveloped virus was found only in neurons. Infected glial cells contained only nonenveloped virus. Neurons in retinal transplants are labeled after PRV injection into the host brain, indicating synaptic connectivity between transplants and degenerated host retinas. This study provides evidence that PRV spreads in the retina as in other parts of the CNS and is useful to outline transplant-host circuitry.

Animals↗

PubMed2004

Optokinetic test to evaluate visual acuity of each eye independently.

A previously described optokinetic testing apparatus [Nat. Neurosci. 5 (2002) 53] was modified to measure vision in each eye separately for evaluation of monocular treatments. This apparatus consists also of a striped rotating drum. Ca. 170 degrees of the drum are illuminated from outside and ca. 190 degrees of the drum move behind a stationary black wall. The rat sits unrestrained in the drum center in a tube so that one eye is unexposed to the rotating stripes. Normal pigmented and retinal degenerate transgenic S334ter-3 rats were tested with the original and the modified apparatus. The usefulness of this method was tested in retinal degenerate rats with retinal transplants in one eye. In retinal degenerate animals, the amount of time (seconds) spent for head-tracking tended to be higher with the original method, possibly due to simultaneous stimulation of both eyes. In rats with retinal transplants, visual responses were significantly preserved in transplanted eyes at late stages of retinal degeneration. In conclusion, contributions from the fellow eye to the optokinetic tracking response can be limited by this testing modification, which is useful for evaluation of treatment effects to one eye.

Age Factors↗

PubMed2004

Vision change after sheet transplant of fetal retina with retinal pigment epithelium to a patient with retinitis pigmentosa.

OBJECTIVE: To report the subjective and objective improvement in vision in a patient with autosomal dominant retinitis pigmentosa after transplantation of a sheet of fetal neural retina together with its retinal pigment epithelium. DESIGN: A sheet of fetal neural retina with its retinal pigment epithelium was transplanted into the subretinal space under the fovea unilaterally in a patient with retinitis pigmentosa with visual acuity of 20/800 in the treated eye. Early Treatment Diabetic Retinopathy Study visual acuity testing, scanning laser ophthalmoscope, tissue typing of the donor and recipient, fluorescein angiography, multifocal electroretinogram, multifocal visually evoked potential, and clinical examination were used. RESULTS: No clinical evidence of rejection was observed. There was no retinal edema or scarring. The transplant sheet lost its pigmentation by 6 months. MAIN OUTCOME MEASURES: A change in visual acuity from 20/800 to 20/400 (7 months), 20/250 (9 months), and 20/160 (1 year) was observed by Early Treatment Diabetic Retinopathy Study visual acuity testing. Independently, scanning laser ophthalmoscope testing at a different institution at 9 months showed a visual acuity of 20/270 at a 40 degrees field of view. CONCLUSION: This study indicates that fetal retina transplanted with its retinal pigment epithelium can survive 1 year without apparent clinical evidence of rejection and show continued improvement in Early Treatment Diabetic Retinopathy Study visual acuity.

Electroretinography↗

PubMed2004

Superior colliculus responses to light - preserved by transplantation in a slow degeneration rat model.

PURPOSE: To determine whether retinal transplantation can preserve visual responses in the superior colliculus (SC) of the S334ter-line-5 rat, a transgenic model for slow photoreceptor degeneration, which is more similar to human retinitis pigmentosa than the fast degeneration line 3 S334ter rat. METHODS: Visual responses to a light flash were recorded in the SC. Rats that had received embryonic day (E) 19-20 fetal retinal sheet transplants at the age of 26-30 days were tested at the ages of 200-254 days. Controls were age-matched rats without surgery and with sham surgery. As a baseline, in no-surgery line-5 rats, the temporal pattern of visual sensitivity loss was evaluated electrophysiologically in the SC from 60 days up to one year of age. RESULTS: In untreated S334ter-line-5 rats, decline in visual sensitivity in the SC was parallel to the photoreceptor loss. At 109 day of age, a relative scotoma developed in the area of the SC corresponding to the nasal retinal region. At 200-254 days of age, the majority of the SC was devoid of any light-driven responses. In contrast, at this time point, transplanted rats with 'good' retinal grafts with normal lamination had visual responses in the caudal region of the SC, the area corresponding topographically to the transplant location in the retina. In these rats, the various parameters of SC responses such as the latency of the onset of the visual response, the response peak amplitude and the consistency of the visual response were significantly different from the control groups (no-surgery, sham surgery, 'poor' transplants) and were more comparable to normal albino rats, however, with a slightly longer latency (70-90 vs. 30-50 msec). CONCLUSIONS: Fetal retinal sheet transplantation showed a long-term rescue effect on visual function in this animal model of slow photoreceptor degeneration.

Animals↗

PubMed2004

Progress in retinal sheet transplantation.

The aim of retinal transplantation is to prevent blindness and to restore eyesight, i.e. to rescue photoreceptors or to replace damaged photoreceptors with the hope of re-establishing neural circuitry. A promising experimental paradigm is the sub-retinal transplantation of sheets of fetal retina, with or without its attached retinal pigment epithelium (RPE), into recipient rats with retinal degeneration. Sheets of fetal retina have already developed their primordial circuitry. Such transplants can develop lamination resembling a normal retina dependent on the presence of healthy RPE either from the host or from the graft. In several retinal degeneration models, transplants have been shown to restore visually evoked responses in an area of the superior colliculus corresponding to the placement of the transplant in the retina. The functional effect of transplants may be due to transplant/host connectivity and/or rescue of host photoreceptors. In summary, sheets of fetal retina can morphologically repair an area of a degenerated retina, and there is evidence to suggest that transplants form synaptic connections with the host and restore visual responses in rats with retinal degeneration.

Animals↗

PubMed2003

Retinal transplantation-induced recovery of retinotectal visual function in a rodent model of retinitis pigmentosa.

PURPOSE: To map the spatiotemporal decline in retinally driven activity in the superior colliculus (SC) of transgenic S334ter-line-3 rats that express a mutated rhodopsin, which causes photoreceptor degeneration. To determine whether transplantation of fetal retinal sheets into the subretinal space of these rats can recover visual activity in the SC. METHODS: A visual stimulus was presented to the eye, and responses were recorded across the SC of untreated S334ter-line-3 rats aged 28 to 288 days. These data were used to draw a map of the developing scotoma. Intact retinal sheets from embryonic day 19 rats were transplanted into the subretinal space of S334ter-line-3 rats between 21 and 28 days of age. Responses to retinal stimulation were mapped in the SC of transplanted and sham control rats 78 to 163 days after surgery. The morphology of the retinas in all groups was examined. RESULTS: Photoreceptor cell loss in untreated rats matched the decline in visual activity in the SC. At 28 days, there was a scotoma in the area of the SC that represents the central retina and, by 63 days, it had enlarged to cover the entire retinal representation. Visual responses were evoked in 64% of rats with retinal transplants. These retinally driven responses were confined to a small, contiguous region of the SC that represents the sector of the retina where the transplant was placed. Visual responses were absent in the SC outside this area in transplant recipients and throughout the SC of untreated and sham control rats. CONCLUSIONS: Transplantation of fetal retinal sheets induced recovery of visual activity in the SC in this model of RP. The mechanisms underlying this functional recovery remain to be resolved, but these results suggest that transplantation should be further explored as a therapy for RP.

Animals↗

PubMed2002

Differential lineage restriction of rat retinal progenitor cells in vitro and in vivo.

To identify and characterize the lineage potential of rat neural retina progenitor cells (NRPCs) in vitro and engrafted into rats with retinal degeneration, NRPCs were isolated from neural retinas of embryonic day 17 Long Evans rats and cultured in serum-free or serum-containing media with fibroblast growth factor 2 and neurotrophin 3. After expansion, cellular differentiation was initiated by the withdrawal of these growth factors. Despite forming primary neurospheres, NRPCs cultured in serum-free medium survived poorly after passage. In contrast, NRPCs cultured in serum-containing medium could be expanded for up to 12 passages and differentiated into glial fibrillary acidic protein-positive glial cells and retina-specific neurons expressing rhodopsin, S-antigen, calbindin, recoverin, and calretinin. For in vivo analysis, passage 1 (P1) undifferentiated NRPCs were labeled with bromodeoxyuridine (BrdU), implanted into the subretinal space of Royal College of Surgeons (RCS) rats, and analyzed immunohistochemically 4 weeks postgrafting. The grafted NRPCs showed extensive glial differentiation, irrespective of their topographic localization. A few BrdU-labeled grafted NRPCs expressed protein kinase C, a marker for bipolar and amacrine interneuron-specific differentiation. Other retina-specific or oligodendrocytic differentiation was not detected in the grafted cells. Although NRPCs are capable of self-renewal and multilineage differentiation in vitro, they developed mostly into glial cells following engraftment into the adult retina. These data suggest that the adult retina retains epigenetic signals that are either restrictive for neuronal differentiation or instructive for glial differentiation. Induction of lineage-specific cell differentiation of engrafted NRPCs to facilitate retinal repair will likely require initiation of specific differentiation in vitro prior to grafting and/or modification of the host environment concomitantly with NRPC grafting.

Animals↗

PubMed2002

Transplanted sheets of human retina and retinal pigment epithelium develop normally in nude rats.

This study investigated whether transplanted sheets of human fetal retina together with its retinal pigment epithelium (RPE) could develop and maintain their cytoarchitecture after long survival times. Transplant recipients were nine albino athymic nu/nu rats with a normal retina. The donor tissue was dissected from fetuses of 12-17 weeks gestational age. Transplants were analyzed at 5-12 months after surgery by light and electron microscopy, and immunohistochemistry with various antibodies specific for rhodopsin, S-antigen, transducin, neurofilament and synaptophysin. In 4 of 11 transplants, the RPE stayed as a monolayer sheet and supported the development of the retinal sheet with a normal lamination, including photoreceptor inner and outer segments. Cones and rods in the organized transplants were labeled with different photoreceptor markers. Inner and outer plexiform layers, containing cone pedicles and rods spherules, were immunoreactive for synaptophysin. As the recipients had a normal retina, transplant/host integration was not expected. However, at the transplant/host interface, there were sometimes areas without glial barriers, and neurofilament-containing processes could be observed crossing between transplant and host. In other, more disorganized transplants, the RPE cells were partially dispersed or clumped together in clusters. Such transplants developed photoreceptors in rosettes, often with inner and outer segments. In conclusion, sheets of human fetal retina transplanted together with its RPE to the subretinal space of nude rats can develop and maintain perfectly laminated transplants after long survival times, indicating the potential of applying cotransplantation to human patients with retinal diseases.

Animals↗

PubMed2002

In vitro isolation and expansion of human retinal progenitor cells.

Human retinal development proceeds with temporal and spatial precision. Although differentiation starts around the beginning of the third month of gestation, the majority of cells in the outer neuroblastic layer of human neural retina are still proliferating, as evidenced by their Ki-67 immunoreactivity. In the present study, the proliferating human retinal progenitor cells (HRPCs) were isolated and expanded in culture. They were capable of dividing for multiple generations (with passage 8, the latest tested) and differentiating to several retinal cell phenotypes. These findings indicate that human retina at the 10th-13th week of gestation harbors progenitor cells that can be maintained and expanded in vitro for multiple generations. The availability of such cells may have important implications with respect to human degenerative retinal diseases, as these HRPCs have the potential to be used therapeutically to replace damaged retinal neurons.

Cell Culture Techniques↗

PubMed2002

Transplantation of intact sheets of fetal neural retina with its retinal pigment epithelium in retinitis pigmentosa patients.

PURPOSE: To show the safety of transplanting sheets of fetal neural retina together with its retinal pigment epithelium (RPE) to patients with retinitis pigmentosa. DESIGN: Interventional case series. METHODS: Sheets of fetal neural retina and RPE were transplanted together into the subretinal space near the fovea unilaterally in the eyes of five patients with retinitis pigmentosa who had only light perception in both eyes. The patients were followed for 6 months. The main outcome measures were tissue typing of both donors and recipients, fluorescein angiography, multifocal electroretinogram (mfERG) testing, and clinical examination. No immunosuppressive medications were given. RESULTS: No evidence of rejection was observed. Up to 6 months there was no evidence of tissue disintegration, retinal edema, or scarring. There was no change in vision both by Snellen acuity and with mfERGs. Growth of the transplant was noted in two of five patients at 6 months vs. 2 weeks. All patients typed were HLA mismatched with donor tissue. CONCLUSIONS: This study indicates that fetal retina can be transplanted together with its RPE and survive for at least 6 months without evidence of rejection. However, no improvements in vision were observed, possibly due to the severe retinal degeneration of the patients.

Adult↗

PubMed2002

Retinal transplantation--advantages of intact fetal sheets.

Retinal transplantation aims to prevent blindness and to restore eyesight, i.e., to rescue photoreceptors or to replace damaged photoreceptors with the hope of reestablishing neural circuitry. Retinal donor tissue has been transplanted as dissociated cells or intact sheets. A promising experimental paradigm is the subretinal transplantation of sheets of fetal retina with or without its attached retinal pigment epithelium (RPE) into recipient rats with retinal degeneration. As long as healthy RPE either from the host or from the graft is present, such transplants can develop lamination resembling a normal retina. Different methods have been used to demonstrate transplant/host connectivity. In two different rat retinal degeneration models, visually evoked responses can be demonstrated in an area of the superior colliculus corresponding to the placement of the transplant in the retina. In summary, sheets of fetal retina can morphologically repair an area of a degenerated retina, and there is evidence to suggest that transplants form synaptic connections with the host and restore visual responses in blind rats.

Animals↗

PubMedDate not supplied

Transplantation of neuroblastic progenitor cells as a sheet preserves and restores retinal function.

Diseases affecting the outer retina are incurable once photoreceptors are lost, and these diseases usually cause retinal pigment epithelium (RPE) dysfunction. However, the inner retina can remain functional for some time, even though retinal remodeling occurs as compensation for photoreceptor loss. If the damaged part can be replaced with neuroblastic progenitor and RPE cells as sheets with a beneficial effect on function, vision loss may be prevented and vision may be restored. This review presents an overview of the research of transplanting sheets of neural retina, with or without its RPE, to the subretinal space. In different animal models of retinal degeneration, retinal transplants can morphologically reconstruct a damaged retina, and restore visual sensitivity. Good morphological integration of transplants with the host retina can occur, whereas other transplants exhibit a glial barrier. Synaptic connections between transplant and host have been indicated by transsynaptic tracing. Retinal transplants can restore and preserve visual responses in a small area of the superior colliculus corresponding to the placement of the transplant in the retina. The beneficial effect of retinal transplantation likely involves two mechanisms: trophic effects, e.g., rescue of host cones; and synaptic connectivity between transplant and host retina.

Animals↗

PubMedDate not supplied

Visual functional effects of constant blue light in a retinal degenerate rat model.

Retinal degenerative conditions increase susceptibility to light damage, but rapid retinal degeneration (RD) models show less susceptibility to cyclic dim light. We investigated whether constant blue light (BL) exposure can eliminate the residual visual responses in a comparatively rapid RD rat model. Pigmented rhodopsin mutant S334ter line-3 rat pups (21 days old) were exposed for 5-6 consecutive days to constant BL. Visual behavior was evaluated with an optokinetic head tracking apparatus. Electrophysiological recordings were made from the superior colliculus (SC). S-antigen, red-green opsin and rhodopsin immunoreactive residual photoreceptors were counted. Following BL exposure, head tracking was significantly reduced at 0.25 cycles degree(-1) in 38-day-old line 3 rats. With a 0.125 cycles degree(-1) stimulus, the head tracking performance of 80-day-old BL rats were similar to that of 220-day-old no-BL-treated line-3 rats. SC recordings also revealed a significant decrease in the residual photoreceptor activity. Histological evaluation showed reduction of the rod population in the central area of the light-damaged retina. Exposure to constant BL considerably reduces the residual visual responses in a rapid degenerating RD rat model.

Age Factors↗