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Thread: Activity of implanted stem cells or OEG

  1. #1
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    Activity of implanted stem cells or OEG

    Dear Wise,

    We have a small technical question: do we know the "lifetime" of an implanted stem cell or OEG cells? When do they begin to differentiate? And when do they become efficient?

    You may have already posted it before but we can't find the reference, that's why I'm asking.

    Thanks,
    Albert

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    Senior Member giambjj's Avatar
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    I posed a similar question some time ago to Dr. Young. He said that implanted cells, nerves, omentum, etc., should be absorbed and have done all they are going to do within 6 months.


    Thanks:

    JJG

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    If no objective progress is observed by the end of six months, does it suggest that the implantation is inefficient? Or shall we wait more than six months to observe objective progress - apart from the recoveries followed by the decompression?

    In the first case - no objective progress six months after surgery - what shall we think about the implantation of olfactive cells / stem cells?

    Tiebreaker: should we consider associating a second therapy, for example embryo cells + laser as done by Shimon Rochkind?

    Albert

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    Senior Member giambjj's Avatar
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    Good question. Better pose it to Dr. Young. Since natural recovery can occur for many years post injury, I don't think thay any one knows for sure.


    JJG

  5. #5
    Albert,

    You have asked a very difficult question because nobody, to my knowledge, has ever done an autopsy on a person who has received an OEG transplant and thereby confirmed the survival and distribution of the transplanted OEG cells. I can only guess at what is happening to the cells in the spinal cord based on animal studies and speculations concerning humans.

    MRI visualization of cell transplants. Dr. Huang slowly injects about 5 µliters of cell suspension into the spinal cord above and below the injury site. One µliter is approximately 1 cubic mm. Clinical MRI scans have a resolution of about 1 cubic mm. Thus, it is possible that the injectate can be seen on MRI shortly after the injection. There may be initially some magnetic resonance difference between the transplanted cells and surrounding spinal cord, due to the difference in cell density in the injectate and the spinal cord. However, the cells should spread out over several days. In rats, transplanted OEG cells migrate at the rate of several mm per day. So, there may be a small spot on the MRI scan where the cells are injected but this should disappear over several days as the cells migrate and integrate into the spinal cord. One should not be able to see the cells after a week and this is apparently the case because I have seen MRI's of spinal cords at several weeks after OEG transplants and there is no sign of the injection sites or the cells in the MRI images of the spinal cord. The spinal cords do not look different from before the OEG transplantation. On the other hand, the University of Florida at Gainsville group were able to use MRI to visualize fetal spinal cords transplanted into the syringomyelic cavities of patients with chronic spinal cord injury. Because they put several such spinal cords into the cavity, the magnetic resonance of the transplants contrasts sharply with the fluid in the cyst, they were able to visualize the translants with MRI.

    Poor survival of OEG cells in acutely injured cord. In animal studies, we know that if we inject neonatal rat OEG cells into the rat spinal cord contusion site shortly after injury, the cells rapidly decliine and are gone within 1-2 weeks. Similar losses of adult OEG cells have been reported by other laboratories from contused spinal cords. The acute inflammatory environment of the injured spinal cord may be deleterious to the transplanted cells. If we inject neonatal OEG cells into rat spinal cord surrounding the injury site, i.e. above and below the injury site, the cells do survive better and longer. During the first week after transplantation, the cells migrated a cm or more from the injection site into surrounding cord but avoided the injury site (or alternatively, they may be migrating into the injury site and dying). By 2 weeks, the OEG cells do migrate into the injury site, perhaps because the inflammation at the injury site is declining. Plant, et al. [1] have reported that delayed transplantation of adult OEG cells into the spinal cord at 1 week after injury not only survive better but reduces cavitation and produces a continuous bridge across the injury site.

    Immune rejection of OEG cells in rats. In our experience, neonatal rat OEG cells transplanted into contused spinal cords shortly after injury are usually gone by 2-4 weeks in rats that have not been immunosuppressed. Note that we are transplanting OEG cells from the same strain of rats (Sprague-Dawley to Sprague-Dawley) but the rats are outbred and therefore are not genetically identical to each other. OEG cells from aborted fetuses are also not genetically identical to person that is receiving the transplant. However, if we treat the rats with daily high-dose cyclosporin A (CyA, 10 mg/kg), transplanted OEG cells not only survived for as long as we looked (14 weeks) but apparently proliferated (made more cells) in the spinal cord. If we stop the daily CyA treatment, the cells are rejected within 2 weeks. We have confirmed that, when OEG cells are rejected from the spinal cord, they are also rejected from other parts of the body, suggesting that it is a systemic immune response. Since CyA suppresses the immune system, the above findings strongly suggest that OEG cells are being rejected by the immune system. This may be in part because we are transplanting the OEG cells into the acutely injured spinal cord, where the blood brain barrier is broken and when many inflammatory and immune cells congregate at the injury site. Some of these inflammatory and immune cells (t-lymphocytes) may initiate the systemic immune response that led to rejection of transplanted OEGs not only in the spinal cord but all over the body. The chronically injured spinal cord is very different in that the blood brain barrier has healed and there are relatively few immune cells such as lymphocytes in the spinal cord.

    Immune rejection of OEG cells in humans. I don't know whether the fetal OEG cells implanted by Dr. Huang are rejected from the spinal cords of humans. There are three major differences between what we do in the rats and what Dr. Huang does in humans. First, he is using fetal OEG cells. The Russians have long maintained that human fetal cells do not express tissue compatability antigens as strongly as adult or neonatal cells and hence are not rejected or are rejected very slowly. Second, Dr. Huang is transplanting OEG cells into people with chronic spinal cord injury (6 months to 16 years after injury), presumably long after the blood brain barrier has healed. He is also injecting the cells above and below the injury site. Thus, the peripheral immune system may not be exposed to the transplanted cells and hence the body does not mount a systemic immune response to the cells. Third, there may be differences between human and rat immune responses. For instance, in the rat, the immune response usually activates and eliminates the cells within 4 weeks. In humans, this immune process may be slower particularly in the central nervous system. As a general rule of thumb, I usually consider one rat week to be equal to one human month. So, it is possible that, even there is immune rejection of the cells, they may survive several months in humans compared to several weeks in rats.

    Methylprednisolone effects on OEG survival and regenerative effects. Nash, et al. [2] found that OEG transplants alone did not improve forelimb recovery in rats with partial lesions of cervical corticospinal tract but treatment with methylprednisolone for 24 hours after the transplant markedly improved regeneration and functional recovery of the rats. We have observed that a single bolus dose of methylprednisolone markedly improve OEG transplant survival in contused spinal cords for several weeks after transplantation. This may be because methylprednisolone is a potent anti-inflammatory drug as well as an immunosuppressant. When I was last in Beijing, Dr. Huang and I talked about giving methylprednisolone alongside OEG transplants. In fact, Mike Kowalski from this site may have been the first person to have received combined methylprednisolone and OEG transplantation (posted in this topic)

    Is long term survival of OEG transplants necessary for their beneficial effects?. Dr. Huang has observed a very surprising early recovery of both motor and sensory function in humans during the first few weeks after transplantation. As mentioned before, this early recovery is too rapid for regeneration or remyelination. We have just completed a detailed study of OEG remyelination in rats and find that the remyelination begins 4 or more weeks after transplantation. So, even in rats, a recovery before 4 weeks would be unlikely to be due to remyelination. On the other hand, we should also realize that our study is in acute spinal cord injury and remyelination may be faster in chronically injured spinal cords. Dr. Huang has been thinking that perhaps the OEG cells are "awakening" residual axons that cross the injury site. He has also gone ahead to try the innovative approach of transplanting OEG a second time after recovery has plateaued. I don't know how many patients have received a second transplant but Dr. Huang has already started doing repeat OEG transplants in some patients. It would be of interest to see the results of the second transplants.

    Regenerative sprouting? I have been veering towards a more radical explanation of OEG mediated early recovery: OEG cells may be promoting sprouting of surviving and regenerating axons that cross the injury site. Most people have some surviving axons crossing the injury site. Based on many years of looking at contused rat spinal cords, I also believe that spinal axons are constantly trying to grow across the injury site, for months (or years) after injury. However, if and when they get across the injury site, regenerating axons may not be able to find neurons to connect to. In other words, spinal axons may be constantly trying to grow across the injury site and some probably are getting through but retract (prune) when they do not connect up with neurons below the injury site. OEG cells release many growth factors that may stimulate surviving and regenerating axons to sprout and that may induce synaptic "plasticity", i.e. encouraging synaptic connections to loosen up and allow sprouting terminals to connect. There is some precedence for the concept of sprouting-induced recovery of function because this may be what IN-1 (the antibody that counteracts Nogo) does. Sprouting can occur rapidly, over a period of days, perhaps even hours. This idea is attractive because it would provide a rational basis of a repeated transplant strategy. If OEG cells are stimulating sprouting and allowing axons to reconnect, this may explain the rapid recovery of segmental function.

    Caveats Please understand that the above is speculative and needs to be confirmed on animal experiments. Until definitive data is available from animal studies, we must guess at how best to proceed with OEG transplantation in the human spinal cord. One of the reasons that I have been urging people to be patient is that new information coming out in the next 6 months may significantly improve the beneficial effects of OEG transplants. For example, in the several months, we should know whether combining OEG transplantation and methylprednisolone is better than OEG alone. Many laboratories are feverishly working on OEG transplants to animal spinal cord injury models. At the recent Society for Neuroscience meeting, I counted over 30 laboratories that are now studying OEG cell transplants. I am expecting significant progress in the coming months. Methylprednisolone may be the tip of the iceberg for drugs that facilitate or enhance the beneficial effects of transplanted OEG cells. For example, there are many anti-inflammatory therapies that can be given for longer term than methylprednisolone. Likewise, many laboratories are starting to combine growth factors with OEG transplants in animal models. It is an exciting time for the field.

    Wise.

    References Cited

    1. Plant GW, Christensen CL, Oudega M and Bunge MB (2003). Delayed transplantation of olfactory ensheathing glia promotes sparing/regeneration of supraspinal axons in the contused adult rat spinal cord. J Neurotrauma. 20: 1-16. The Chambers Family Electron Microscopy Laboratory, The Miami Project To Cure Paralysis, Miami, Florida, USA. gplant@anhb.uwa.edu.au. The aim of this study was to determine the preferred time and environment for transplantation of olfactory ensheathing glia (OEG) into the moderately contused adult rat thoracic spinal cord. Purified OEG were suspended in culture medium with or without fibrinogen and injected into the contused cord segment at 30 min or 7 days after injury. Control animals received a contusion injury only or injection of only medium 7 days after contusion. The effects on axonal sparing/regeneration and functional recovery were evaluated 8 weeks after injury. The grafts largely filled the lesion site, reducing cavitation, and appeared continuous with the spinal nervous tissue. Whereas in 7d/medium only animals, 54% of spinal tissue within a 2.5-mm-long segment of cord centered at the injury site was spared, significantly more tissue was spared in 0 d/OEG-medium (73%), 0 d/OEG-fibrin (66%), 7 d/OEG-medium (70%), and 7 d/OEG-fibrin (68%) grafted animals. Compared with controls, the grafted animals exhibited more serotonergic axons within the transplant, the surrounding white matter, and the spinal cord up to at least 20 mm caudal to the graft. Retrograde tracing revealed that all but the 0 d/OEG-fibrin graft promoted sparing/regeneration of supraspinal axons compared with controls. Overall, the 7 d/OEG-medium group resulted in the best response, with twice as many labeled neurons in the brain compared with 7 d/medium only controls. Of the labeled neurons, 68% were located in the reticular formation, and 4% in the red, 4% in the raphe, and 5% in the vestibular nuclei. Hindlimb performance was modestly but significantly improved in the 7 d/OEG-medium group. Our results demonstrate that transplantation of OEG into the moderately contused adult rat thoracic spinal cord promotes sparing/regeneration of supraspinal axons and that 7 d transplantation is more effective than acute transplantation of OEG. Our results have relevant implications for future surgical repair strategies of the contused spinal cord.

    2. Nash HH, Borke RC and Anders JJ (2002). Ensheathing cells and methylprednisolone promote axonal regeneration and functional recovery in the lesioned adult rat spinal cord. J Neurosci. 22: 7111-20. Neuroscience Program, Department of Anatomy, Physiology, and Genetics, F. Edward Hebert School of Medicine, Uniformed Services University of the Health Sciences, Bethesda, Maryland 20814-4799, USA. hollyhnash@yahoo.com. Axons fail to regenerate after spinal cord injury (SCI) in adult mammals, leading to permanent loss of function. After SCI, ensheathing cells (ECs) promote recovery in animal models, whereas methylprednisolone (MP) promotes neurological recovery in humans. In this study, the effectiveness of combining ECs and MP after SCI was investigated for the first time. After lesioning the corticospinal tract in adult rats, ECs were transplanted into the lesion, and MP was administered for 24 hr. At 6 weeks after injury, functional recovery was assessed by measuring successful performance of directed forepaw reaching (DFR), expressed as percentages. Axonal regeneration was analyzed by counting the number of corticospinal axons, anterogradely labeled with biotin dextran tetramethylrhodamine, caudal to the lesion. Lesioned control rats, receiving either no treatment or vehicle, had abortive axonal regrowth (1 mm) and poor DFR success (38 and 42%, respectively). Compared with controls, MP-treated rats had significantly more axons 7 mm caudal to the lesion, and DFR performance was significantly improved (57%). Rats that received ECs in combination with MP had significantly more axons than all other lesioned rats up to 13 mm. Successful DFR performance was significantly higher in rats with EC transplants, both without (72%) and with (78%) MP, compared with other lesioned rats. These data confirm previous reports that ECs promote axonal regeneration and functional recovery after spinal cord lesions. In addition, this research provides evidence that, when used in combination, MP and ECs improve axonal regrowth up to 13 mm caudal to the lesion at 6 weeks after injury.

    [This message was edited by Wise Young on 12-03-03 at 01:39 PM.]

  6. #6
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    Dear Dr. Young -

    I know you're totally busy, but I've been waiting at the edge of my seat for your convention update summary!!! Just a gentle reminder .

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    Wise,
    I'll have your post translated to understand it fully and will answer you soon.
    Albert

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    Hi Albert. I posted earlier, but I see that it didn't go through. I will try again! When I was in Portugal I met with some of Dr. Lima's patients that had gone before me and they claim to still be seeing some return after 6 months. Some even said they noticed things as long as a year out of surgery. I do not know though how long they were post injury.
    We are going to France in June for my sister inlaws wedding. I was hoping to possibly be able to put some time aside to come to your clinic. I know summer is a busy time, so I will try to let you know in advance. Tell everyone hello for me. Joy

  9. #9
    ip, sorry. I have been so busy. Wise.

  10. #10
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    Thanks Dr. Young. I'm glad you're busy . Your progress means less time for us in these stupid chairs.

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