Stem Cell Terminology

There was a recent article that I found difficult to read without looking up many terms. It seemed worthwhile to use it as a learning exercise. Here’s the original article (an abstract):

2017-06-30: “Human Umbilical Cord Mesenchymal Stem Cells Protect Against SCA3 by Modulating the Level of 70 kD Heat Shock Protein

Here it is broken down:

Spinocerebellar ataxia 3 (SCA3), which is a progressive neurodegenerative disease, is currently incurable. Emerging studies have reported that human umbilical cord mesenchymal stem cells (HUC-MSCs) transplantation could be a promising therapeutic strategy for cerebellar ataxias.

The significance of the umbilical cord is the presence of cord blood, which contains primitive stem cells. The donor need not be the recipient, so no, your mother need not have banked your umbilical cord. The main alternative sources for stem cells are peripheral blood (deoxygenated, venous blood) and bone marrow, though it’s unclear to me whether this paper is saying there’s something unique about HUC-MSCs that would be absent from other types of stem cells.

Note that neural stem cells (NSCs) are generated in the embryonic stage, and if neurons in the brain die, then they cannot be replaced (except for a few rare exceptions). Stem cells transplanted into the brain will not cause them to differentiate into replacement neurons.

Mesenchymal: relating to embryonic tissue used in the umbilical cord, the vitreous (gel-like substance resembling glass) of the eyeball, etc.

Mesenchymal stem cells: a class of multipotent adult stem cells that generates connective tissues (including cartilage, tendons, and bone).

Transplant: move or transfer something to another place or situation, typically with some effort or upheaval.

Why the word transplantation here? Unless we are talking about maternal twins or using your own stem cells, stem cells of the donor have DNA that is different from the recipient, so there is kind of an implied upheaval.

However, few studies have evaluated the effects of HUC-MSCs on SCA3 transgenic mouse.

We know transgenic mice from the last time I created a page like this. But human stem cells transplanted into mice?? That is odd.

Thus, we investigated the effects of HUC-MSCs on SCA3 mice and the underlying mechanisms in this study. SCA3 transgenic mice received systematic administration of 2 × 106 HUC-MSCs once per week for 12 continuous weeks.

Not sure of the units (106). I’ll assume the 2x means twice in one day.

I wonder what administration method was used (ICBI? CVC (doubtful in mice)?).

Motor coordination was measured blindly by open field tests and footprint tests.

I’ll assume the meaning is what is intuitively obvious.

Immunohistochemistry [staining] and Nissl staining were applied to detect neuropathological alternations.

Staining: Stains and dyes are used to highlight structures in biological tissues for viewing, often with the aid of microscopes.

Immunohistochemistry (IHC): the roots are immuno, in reference to antibodies used in the procedure, and histo, meaning tissue. IHC staining is used in the diagnosis of abnormal cells.

Nissl staining: a nucleic acid (related to DNA/RNA) staining method used on nervous tissue sections. Named after Franz Nissl.

Neuropathology is the study of disease of nervous system tissue, usually in the form of either small surgical biopsies or whole-body autopsies.

Alternation: the repeated occurrence of two things in turn. I wonder if this is a misspelling of alteration.

Neurotrophic factors in the cerebellum were assessed by ELISA.

Trophic: relating to feeding and nutrition.

Neurotrophic: relating to the growth of nervous tissue.

Neurotrophic factors (NTFs): a family of biomolecules that support the growth and survival of neurons.

ELISA: Enzyme-linked immunosorbent assay (ELISA) is a test that uses antibodies and color change to identify a substance.

We used western blotting to detect the alternations of heat shock protein 70 (HSP70), IGF-1, mutant ataxin-3, and apoptosis-associated proteins.

I wonder, again, if alternation is misspelled (or misused).

Blot: a method of transferring proteins, DNA, or RNA onto a carrier so they can be visualized by colorant staining.

Western blot: The western blot (sometimes called the protein immunoblot) is a technique for detecting specific proteins in a tissue sample. (The name is a pun, keying off a related method named after Edwin Southern.)

Heat shock: the effect of subjecting a cell to a higher temperature than that of the ideal body temperature. Heat shock can cause proteins (created by the cells) to misfold, aggregate, and/or entangle.

Heat shock protein (HSP): a family of (protective?) proteins that are produced by cells in response to exposure to stressful conditions.

Heat shock protein 70 (HSP70): a family of ubiquitously expressed heat shock proteins. They are an important part of the cell’s machinery for protein folding and help to protect cells from stress. The 70 comes from weighing 70 kilodaltons.

Insulin-like growth factor 1 (IGF-1): a hormone similar in molecular structure to insulin. It plays an important role in childhood growth and continues to have anabolic effects in adults (relating to the synthesis of complex molecules for the storage of energy).

Mutant ataxin-3: the underlying protein problem that SCA3 causes!

Apoptosis: programmed (i.e., planned, expected, necessary; thereby efficient and cleaned up) cell death. As opposed to necrosis, which is traumatic cell path.

Apoptosis-associated proteins: proteins found to be present during the apoptosis process.

Tunel staining was also used to detect apoptosis of affected cells.

TUNEL: stands for TdT (Terminal Deoxynucleotidyl Transferase)-Mediated dUTP (2′-Deoxyuridine 5′-Triphosphate) Nick-End Labeling. It is a method for detecting DNA fragmentation during apoptosis.

This wins the prize for ugliest acronym expansion ever.

The distribution and differentiation of HUC-MSCs were determined by immunofluorescence.

Differentiation: stem cells changing into a specific cell type.

Immunofluorescence: a technique that uses the specificity of antibodies to their antigen to target fluorescent dyes to specific biomolecule targets within a cell and therefore allows visualization of the distribution of the target molecule through the sample.

Our results exhibited that HUC-MSCs transplantation significantly alleviated motor impairments, corresponding to a reduction of cerebellar atrophy, preservation of neurons, decreased expression of mutant ataxin-3, and increased expression of HSP70.

Understandable now.

Implanted HUC-MSCs were mainly distributed in the cerebellum and pons with no obvious differentiation, and the expressions of IGF-1, VEGF, and NGF in the cerebellum were significantly elevated.

Stem cells, administered in an unspecified way, ended up both in the cerebellum and pons. In other words, targeting wasn’t specific to the cerebellum either by choice or by outcome.

Vascular endothelial growth factor (VEGF): a signal protein produced by cells that stimulates the formation of blood vessels. It is part of the system that restores the oxygen supply to tissues when blood circulation is inadequate such as in hypoxic conditions.

Nerve growth factor (NGF): protein-like molecules primarily involved in the regulation of growth, maintenance, proliferation, and survival of certain target neurons.

Elevated? Apparently, boosting these levels is good.

Furthermore, with the use of HSP70 analogy quercetin injection, it demonstrated that HSP70 is involved in mutant ataxin-3 reduction.

Biological analogy: normally, this would refer to a commonality between transgenic mice and humans.

Quercetin: an antioxidant, readily available in supplement form. I take it as a natural anti-inflammatory.

I wonder about the word analogy here. It could be that something else was meant. The web indicates that quercitin inhibits HSP expression.

These results showed that HUC-MSCs implantation is a potential treatment for SCA3, likely through upregulating the IGF-1/HSP70 pathway and subsequently inhibiting mutant ataxin-3 toxicity.

Upregulation: essentially an increase.

Back to the title:

Human Umbilical Cord Mesenchymal Stem Cells Protect Against SCA3 by Modulating the Level of 70 kD Heat Shock Protein

Translation: Stem cells protect against SCA3 by triggering an increase in protein protectants.

In my opinion, this whole song and dance is still disease amelioration at best.

Addendum—2018 trial

I’m not a proponent of stem cells and SCA; I’m just trying to keep up with what’s going on. There is a phase 2 trial beginning 2018 January, planned for two years, sponsored by a Chinese company called SCLnow Biotechnologies, who has a very low-quality website.

Clearer Thoughts on a Cure

Some of the spinocerebellar ataxias, including SCA3, are genetically similar to Huntington’s disease (HD) in that they involve CAG repeats on a chromosome. I’ve found the HD community to be more informed and practical than the SCA community, where:

  • There’s not a misinformation campaign against accurately conveying disease prevalence, as there is for SCA.
    • Sadly, they do have their own special brand of nonsense, around a meaningless and fabricated at-risk prevalence number of 200,000.
  • They’re more up-front about the impossibility of a gene-based cure, which I believe is a taboo subject in the world of SCA. It took me a few decades to unravel this basic notion, because there’s so much pressure to deny it in the world of SCA. That is the topic of this article.

Background

Blog content consolidated here.

Curability

Someone with SCA cannot be cured in the sense of ridding the genetic / neuronal / chromosomal problem from their body and/or brain, as the body is built on trillions of copies of the same genome. In some circles, the phrase disease-modifying is replacing the word curing since the disease cannot literally be cured but will possibly be modifiable to a fractional degree in the future. There are thousands of genetic diseases; not one has been “cured.”

For those with an SCA defect, there are two main cases to consider:

1. Being asymptomatic: The goal in this case is to live one’s life with the defect but never become symptomatic. This is the most desirable outcome for those with the defect.

There are two theoretical possibilities here for achieving that: (1) gene silencing—correcting the toxic protein as it’s created, and (2) counteracting the toxic protein that gets created before it damages the cerebellum.

There are a few glimmers of possible drug therapies, but it’s unwise to make long-term predictions. Given the unknown time horizon (decades?) and unknown effectiveness, limitations, and side-effects of future drugs, there’s the possibility that no solution like this will be developed.

2. Being symptomatic: The goal in this case is to stop or at least slow the cerebellar degeneration. I think this is the likeliest achievable case, still decades out, and barely desirable. The desirability is low because symptoms won’t likely improve: if cerebellar degeneration can truly be halted (unlikely; slowed is more realistic), then symptoms could be frozen in time for the rest of one’s life and begin to interact with the aging process.

If slowing degeneration is the best case, then case 2 really equals case 1. Having the genetic defect but no disease symptoms means the symptoms are pending, unless you die first.

Interestingly, in 2018, there is an effort to separate #1 and #2 and classify #1 as being “clinically ready,” which by implication seems to classify #2 as clinically hopeless.

Notes

There might end up being just one drug therapy approach that covers both cases above. Picture a world where everyone with the SCA(3) defect takes the same drug whether they have symptoms or not: those without symptoms hope not to develop them, and those with symptoms are in the nebulous territory of hoping not to worsen.

Symptoms are not an exact reflection of the cerebellar degeneration that has occurred (i.e., cell damage vs. cell death). External symptoms might occur years after cerebellar degeneration has begun. To remain symptom free, I’d guess that one with the defect would need to begin diligent drug therapy in early childhood or infancy, perhaps in utero. (Case in point: nusinersen; a more explicit example.) If one waits until there are external symptoms, the amount of and permanence of cerebellar damage, and the momentum of the damage, might be insurmountable, especially as the aging process marches forward.

Someone taking medication to thwart symptoms in themselves can still pass on the defect to their offspring, via their sperm or ova. To avoid passing on the defect, the prevention ideas (below) must still be followed. As disappointing as it is, I imagine a future world where those with genetic diseases can mask them with a lifetime of medication, all the while passing on the defect to future generations, ensuring that the diseases are never eradicated.

The conundrum becomes this: slowing the disease to beyond one’s lifetime could be just as good as stopping it altogether, but how do we get there from here? Therapies will need to evolve for decades to achieve this result, yet the early, undesirable therapeutic stages will still cost millions of USD dollars yearly, making them even more undesirable.

Prevention

The only theoretically perfect long-term solution to SCA is prevention. “Perfect” means babies are born free of the defect. Anyone born with the defect must deal with it as they age. If they’ve been tested and understand their situation, I think they have an obligation to humanity to take evasive maneuvers in the procreation process, but some are strongly opposed to this idea.

Since the early to mid-1990s when DNA testing became available, we have had everything needed to rid the world of various SCA diseases in one generation, while still allowing for offspring. Prevention can only revolve around pregnancy (avoiding or modifying it). There is no other place in the life cycle to apply principles of prevention.

Here’s what’s working for us if you have an SCA defect and know it:

  • You can choose not to have offspring.
  • Genetic testing of blastocysts, embryos, fetuses, children, and adults is possible, at any time (asymptomatic or symptomatic). In the context of prevention, this is important so that testing can be done at the fertilization or fetal stage of one’s potential offspring.
  • Amniocentesis followed by possible fetal abortion.
  • In vitro fertilization (IVF) with preimplantation genetic diagnosis (PGD), to avoid possible abortions.

Here’s what’s working against us :

  • The expense of genetic testing put it (and the items below) out of reach for many.
  • Possible religious opposition to fetal abortion.
  • The expense and nontrivial nature of IVF with PGD (USD 20,000 or so).
    • Possible religious opposition to destroying embryos, both the runoff (from IVF) and the rejected embryos (from PGD).
    • This technique isn’t guaranteed to result in a successful pregnancy. No technique is guaranteed.
  • Some are opposed to applying any technology to the procreation process. Some are opposed to using DNA testing as a tool.
  • If you have kids early, and don’t know of the defect, you won’t know any of this. Factors to consider here are if you don’t know who at least one of your parents was, or if the parent you inherited the defect from died without an SCA diagnosis.

It’s ironic that if drug therapies are developed that allow one to have SCA but not develop symptoms, I’d say it’s human nature to keep having kids without safeguards, thereby allowing the defect to exist in the world forever. I don’t think prevention will be achieved.

Does the work being done have a long-term vision of eradicating the diseases from future generations (no), or just looking at snake-oil impact on the current generation (yes)? I’d say we are quite literally gearing up for humanity to have these diseases into the future with unwavering prevalence, possibly minimizing their impact on those willing to be tested and take medication.

SCA pipe dreams

The most famous SCA pipe dream is anything involving stem cells. Stem cells will never be a part of a perfect SCA solution—maybe only aiding, indirectly, as an in vitro platform for drug testing, i.e., for growing things to test on. Maybe in the future (perhaps even now), stem cells safely injected directly into the brain will offer temporary improvements, but they will never prevent or eradicate the SCA problem in an individual. The new cells would only supplant glial cells and not preventatively replace portions (neurons) of one’s functioning cerebellum.

The up-and-coming SCA pipe dream is that single-cell CRISPR (i.e., gene editing) can help with it. CRISPR without a mechanism of delivery into mature organisms works at the zygote stage (even better at the fertilization stage), maybe getting into the blastocystic or embryonic stages. To fix SCA in an adult, a delivery mechanism is needed to fix the billions of neurons in a mature cerebellum.

Does single-cell CRISPR offer us with SCA anything over IVF with PGD, which has been available for 20+ years? No—it would still be used in conjunction with IVF and involve the discarding of unused embryos. IVF with PGD (available for 20+ years) can be used to discard zygotes that are determined to have a genetic defect, whereas single-cell CRISPR can theoretically fix a defective genome before cell replication begins.

I think that the latest fervor over antisense oligonucleotides (ASOs—a form of gene silencing, not gene editing) will be overtaken with CRISPR fever, because even though neither offers a perfect solution, CRISPR will be seen as better than ASOs because it’s more permanent—modifying the DNA in the genome, rather than leaving the defect in the DNA and affecting only the messenger RNA.

My follow-up on CRISPR.