The authors have no acknowledgements.

The study protocol was approved by the Ethics Committee of the Low Vision Academy and all subjects signed a written consent in accordance with the Helsinki Declaration. This research study has received ethical approval from both Loughborough and Sheffield Universities.
NOTE: The inclusion and exclusion criteria of dry age-related macular degeneration patients to receive suprachoroidal autologous graft by Limoli Retinal Restoration Technique (LRRT) is described in Table 1.
...

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G., Vingolo, E. M., Morales, M. U., Nebbioso, M., Limoli, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Preliminary+Study+on+Electrophysiological+Changes+After+Cellular+Autograft+in+Age-Related+Macular+Degeneration.">Preliminary Study on Electrophysiological Changes After Cellular Autograft in Age-Related Macular Degeneration.</a> Medicine. 93 (29), 355 (2014).</li><li>Tischler, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Platelet+rich+plasma:+The+use+of+autologous+growth+factors+to+enhance+bone+and+soft+tissue+grafts.">Platelet rich plasma: The use of autologous growth factors to enhance bone and soft tissue grafts.</a> N. Y. State Dent. J. 68, 22 (2002).</li><li>Zuk, P. 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The primary purpose of this study was to evaluate whether the suprachoroidal graft of adipocytes, ADSCs in SVF, and PRP could improve VA and retinal sensitivity in dry AMD-affected eyes over time. Another main objective was to demonstrate possible therapeutic effects of these cells, based on the recent literature, since several preclinical studies have suggested that GF-based therapy could be useful for patient care in several diseases.
In fact, some studies have shown that autologous human in...

Cell therapy, consisting of the systemic or local injection of stem/progenitor cells in the injured area to treat multiple chronic disorders, has drawn close attention in the last decade1. Since the 1990s, growth factors (GFs) have been studied for their potentially therapeutic role in retinal atrophy2. In fact, many human cells can produce GFs, which are specific proteins that are able to block or slow down apoptosis, i.e., the programmed death of cells3.
It is known that dry age-related macular degeneration (AMD) is an atrophic retinal disease where gradual and irreversible cell death involves injury to the photoreceptor layer and, consequently, the loss of central visual function4. AMD is the leading cause of blindness in people over 55 years of age in developed countries and accounts for 80% of all macular degenerations, which lack an effective treatment to date.
Several studies have shown that there are various sources from which autologous GFs can be obtained. These comprise different cell types, including adipose stromal cells derived from orbital fat, platelets derived from platelet-rich plasma (PRP), and adipose-derived stem cells (ADSCs) included in the stromal vascular fraction (SVF) of adipose tissue5,6,7. The current GF set ensures retinal neuroenhancement, and research conducted by Filatov, Meduri, Pelaez, and Limoli has demonstrated that autologous fat transplantation (AFT) is effective8,9,10.
Moreover, a prior study showed significant improvements in electroretinogram (ERG) data, recorded post suprachoroidal autologous graft, in dry AMD-affected eyes11. The surgically grafted tissue in the suprachoroidal space modulated the paracrine secretion of retinal cells, delaying their apoptosis6,7,12. Considering outer nuclear layer thickness, the histological examination of the retina of guinea pigs has shown that GFs could have a trophic effect on the retina. Therefore, the direct or indirect use of GFs can potentially bring therapeutic benefits through a balanced relation between molecular inducers and inhibitors6,7,12.
The purpose of this method is to assess whether the suprachoroidal graft of adipocytes, ADSCs in SVF and PRP can improve best corrected visual acuity (BCVA) and microperimetry (MY) responses in dry AMD-affected eyes. This study aims to demonstrate the therapeutic effect of autograft on the basis of its GF production, according to the cited literature6,7,12,13.

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	<tbody>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Blunt cannula, 3 mm.&#160;</td>
			<td style="width:179px;">Mentor, Santa Barbara, CA.</td>
			<td style="width:119px;"></td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Luer-LokTM syringe.&#160;</td>
			<td style="width:179px;">BD Biosciences, Franklin Lakes, NJ.</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Regen-BCT tube.&#160;</td>
			<td style="width:179px;">RegenKit; RegenLab, Le Mont-sur-Lausanne, CH.</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">Centrifuge</td>
			<td style="width:179px;">&#160;RegenPRP Centri. RegenLab, Le Mont-sur-Lausanne, CH.</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">BD Venflon Pro Safety 22G x 1.00 inch (0.9 mm x 25 mm).&#160;</td>
			<td style="width:179px;">BD Biosciences, Franklin Lakes, NJ.</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">SPSS Statistics Version 19.0</td>
			<td style="width:179px;">IBM Corp., Armonk, NY, USA.</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Confocal scanning laser ophthalmoscope&#160;</td>
			<td style="width:179px;">Nidek Inc, Fremont, CA</td>
			<td style="width:119px;">Nidek F10&#160;</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Cirrus 5000 Spectral Domain-Optical Coherence Tomography</td>
			<td style="width:179px;">Carl Zeiss Meditec AG, Jena, Germany</td>
			<td style="width:119px;">&#160;SD-OCT&#160;</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Maia 100809 Microperimetry&#160;</td>
			<td style="width:179px;">CenterVue S.p.A., Padua, Italy</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Ocular electrophysiology electromedical system,</td>
			<td style="width:179px;">C.S.O., S.r.l., Scandicci, Italy</td>
			<td style="width:119px;"></td>
			<td>&#160;Retimax for ERG&#160;</td>
		</tr>
	</tbody>
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regenerative therapy by suprachoroidal cell autograft in dry age-related macular degeneration: preliminary in vivo report

This study is aimed at examining whether a suprachoroidal graft of autologous cells can improve best corrected visual acuity (BCVA) and responses to microperimetry (MY) in eyes affected by dry Age-related Macular Degeneration (AMD) over time through the production and secretion of growth factors (GFs) on surrounding tissue. Patients were randomly assigned to each study group. All patients were diagnosed with dry AMD and with BCVA equal to or greater than 1 logarithm of the minimum angle of resolution (logMAR). A suprachoroidal autologous graft by Limoli Retinal Restoration Technique (LRRT) was carried out on group A, which included 11 eyes from 11 patients. The technique was performed by implanting adipocytes, adipose-derived stem cells obtained from the stromal vascular fraction, and platelets from platelet-rich plasma in the suprachoroidal space. Conversely, group B, including 14 eyes of 14 patients, was used as a control group. For each patient, diagnosis was verified by confocal scanning laser ophthalmoscope and spectral domain-optical coherence tomography (SD-OCT). In group A, BCVA improved by 0.581 to 0.504 at 90 days and to 0.376 logMAR at 180 days (+32.20%) postoperatively. Furthermore, MY test increased by 11.44 dB to 12.59 dB at 180 days. The different cell types grafted behind the choroid were able to ensure constant GF secretion in the choroidal flow. Consequently, the results indicate that visual acuity (VA) in the grafted group can increase more than in the control group after six months.

Using the procedure presented here, two groups of dry AMD-affected patients, with BCVA equal to or greater than 1 logarithm of the minimum angle of resolution (logMAR), were enrolled in the study. Group A, including 11 eyes of 11 patients, received suprachoroidal autologous graft by Limoli Retinal Restoration Technique (LRRT), whereas group B, including 14 eyes of 14 patients, was used as a control group.
Student&#39;s t-test an...

Watch this Scientific Journal Video about Regenerative Therapy by Suprachoroidal Cell Autograft in Dry Age-related Macular Degeneration: Preliminary In Vivo Report at JoVE.com

Regenerative Therapy by Suprachoroidal Cell Autograft in Dry Age-related Macular Degeneration: Preliminary In Vivo Report

The goal of this study is to assess whether the suprachoroidal graft of adipose-derived stem cells included in the stromal vascular fraction and platelets derived from platelet-rich plasma by the Limoli Retinal Restoration Technique can improve visual acuity and retinal sensitivity responses in eyes affected by dry age-related macular degeneration.

Regenerative Therapy by Suprachoroidal Cell Autograft in Dry Age-related Macular Degeneration: Preliminary In Vivo Report

This study is aimed at examining whether a suprachoroidal graft of autologous cells can improve best corrected visual acuity (BCVA) and responses to ...

The overall goal of this surgical intervention is to introduce a retinal neuro enhancement that slows retinal cell apoptosis in order to improve visual performance and rehabilitation of low vision patients. This method can help answer key questions in the field of low vision caused by retinal atrophic degeneration, such as a dry AMD or neural optic atrophy. It will improve vision in both the natural setting and vision with an assisted magnifying device.These retinal restoration promote the vascular pedicle fat increasement with the underlying colloid. and it enhances the vascularization of the pedicle, improving the pedicle's health and survival. This therapy also works cause it gives a regenerative startup to all the retinal elements, through the participation of growth factors.Although this technique was developed for slowing retinal degeneration interry, it can be applied also in adult tissues or organ systems where therapy is needed. To begin, use a 22-gauge needle to collect eight milliliters of peripheral blood, in order to collect the platelet-rich plasma. Use a sodium citrate vacuum tube with separator gel, that promotes the separation between cellular components and plasma.Centrifuge the tube of blood for five minutes at 1500g, and 20 degrees Celsius. The blood will separate into three layers. Aspirate and dispose of the top layer, consisting of platelet-poor plasma.Then, collect and save the middle layer, consisting of platelet-rich plasma. Anesthetize the macular degeneration patient for the Limoli retinal restoration in order to harvest the fat tissue. Harvest adipose tissue from the abdominal subcutaneous layer.Collect about 10 milliliters of tissue using a three millimeter blunt cannula, connected to a locking syringe. Use the Lawrence and Coleman technique. Then, separate the stromal vascular fraction of fat tissue by spinning down the sample for five minutes at 1500 and 20 degrees Celsius.The stromal vascular fraction, located just above the blood layer, is very rich in adipose-derived stem cells. Not more than one milliliter is needed for the autograft. Finally, compose the autograft, loading 0.5 to 0.6 milliliters of stromal vascular fraction, 0.2 to 0.3 milliliters of platelet-rich plasma, and 0.1 milliliters of hyaluronic acid into a delivery tube.Blend it by means of two syringes and a three-way stopcock. Angle the tube of the cannula up at five millimeters in order to facilitate the injection of the autograft into the sub-scleral space. Before beginning the surgery, mark the eye that is undertaking the treatment while the patient is in a seated position, to delimitate the working area, avoiding the physiological cyclo-torsion effect of the eye.Begin the restorative surgery by first anchoring the sclera with a 6-0 silk suture near the inferior temporal limbus. Sclera anchoring is a critical step. The needle must enter the limbus not too deeply, not too superficially.If you perforate the eye, it will deflate. If the thread is an oketa, the eye will not be anchored. What a stress!Next, using 5.5 inch Westcott tenotomy curved scissors, open the sub-conjunctival and sub-Tenonian space, 11 millimeters from the inferior temporal limbus. Therein, insert the Limoli basial conjunctival retractor to make a scleral surgical field. Now, using a five millimeter crescent knife, angled bevel up, pre-cut a flap within the inferior temporal quadrant of the sclera, eight millimeters from the limbus.Make the flap hinge radial, and to the left of the surgeon. Next, use the crescent knife, angled bevel up, to open a deep scleral door about five millimeters wide, with a radial hinge in the inferior temporal quadrant, at eight millimeters from the limbus. Cut very slowly, tangentially to the ocular surface.Do not penetrate the choroidal space. The ideal depth is achieved when the bluish color of the underlying choroid appears, and the off-white scleral traces become less visible. At this stage, the choroid is like a thin ice floe.If the Sclerectomy is not deep enough, gram factors produced by implanted cells will not penetrate as desired. If the Sclerotomy is too deep, it risks the choroid perforation, leading to hemorrhage, or bulk perforation, followed by a retinal damage. Now collect some orbital fat.Create a gap by removing a little operculum in the distal part of the flap, in order to facilitate blood circulation in the subsequent supra-choroidal autograft. Then, using ophthalmological forceps, extract the orbital fat from a gap above the inferior oblique muscle. Take note of small bleedings from the pedicle.These are indicative of a good vascularization, important to the survival of the autograft. Orbital fat surgery is another critical step. We open orbital space without scissor, in order to prevent bleeding or muscle cuts.Then, we isolate fat pedicle before anchoring it to the sclera. Next, gently place the autologous fat pedicle on the choroidal bed, and suture it closed along the proximal edge of the door, using choroidal 6-0 polyglactin fiber. Next, suture the scleral flap to avoid compression on the fat pedicle, or on its nutrient vessels.Now, into the stroma of the fat pedicle, infiltrate one milliliter of platelet-rich plasma gel, using a 30-gauge angled cannula. Next, prepare the sides of the conjunctiva for suturing by removing the retractor. Then suture the conjunctiva using 7-0 polyglactin fiber, but do not close it.Leave enough space to insert a small flexible tube into the scleral pocket. Now, insert the tube into the scleral pocket and inject the autograft into the sub-scleral space. Finally, inject the sub-conjunctival hybrid composed of platelet-rich plasma, antibiotics, and steroids.For the next three days, provide systemic antibiotic therapy, and for the next 15 to 20 days, provide eyedrops containing both an antibiotic and a steroid. Patients with dry, age-related macular degeneration were divided into two groups. Each had a best correct visual acuity score of at least one on the BAM log scale.Patients in the control group underwent therapy based on antioxidant supplements and visual rehabilitation when necessary, whereas the experimental group underwent the LRRT. Diagnosis of the results was carried out using a confocal scanning laser ophthalmoscope, and spectral domain optical coherence tomography. After 90 days, the BAM log score of the treatment group improved, and after 180 days, it further improved.Furthermore, the microperimitry test score for LRRT-treated patients increased from an average of 11.44 decibels to an average of 12.59 decibels. Overall, these results suggest that the different cell types grafted behind the choroid were able to ensure constant growth factor secretion into the choroidal flow. And then consequently, the visual acuity of the grafted group increased more than in the control group.After watching this video, you should have a good understanding for a potential therapy for cell apoptosis using a surgery that promotes interactions between growth factors and residual retinal cells. Once mastered, this technique can be done in 13 minutes per eye, if it is performed properly. While attempting this procedure, it's important to tread carefully during scleral anchoring, and while making the deep sclerotomy.Otherwise, this technique is relatively simple and safe. After its development, this technique paved the way for researchers in the field of retinal restoration in order to explore the possibility to stop or slow down the neuro-retinal atrophic degeneration and to improve its visual performance.

regenerative-therapy-suprachoroidal-cell-autograft-dry-age-related

Research

JoVE Journal

Medicine

10.0K vues.  Low Vision Research Centre of Milan. The overall goal of this surgical intervention is to introduce a retinal neuro enhancement that slows retinal cell apoptosis in order to improve visual performance and rehabilitation of low vision patients. This method can help answer key questions in the field of low vision caused by retinal atrophic degeneration, such as a dry AMD or neural optic atrophy. It will improve vision in both the natural setting and vision with an assisted magnifying device.These retinal restoration promote...