New ALS Research Involves TDP-43, Motor Neurons, Other Findings

TDP-43 Regulates Expression of SOD1
Reducing expression of TDP-43 increases the level of SOD1, according to new research funded by The ALS Association.  The finding highlights a previously unknown molecular link between two genetic causes of ALS and may lead to a better understanding of disease pathogenesis.  Mutations in the TDP-43 gene are a rare cause of familial ALS, and the TDP-43 protein is found in protein aggregates in most forms of ALS, both familial and sporadic.  The normal TDP-43 protein helps process RNA in the cell.  It is not yet known how mutations in TDP-43 cause ALS.

The authors employed a reporter assay in which SOD1 protein was linked to part of an enzyme; when it joined to the other part of the enzyme (supplied in excess), it emitted light proportional in brightness to the amount of SOD1 present.  The authors then screened a whole-genome RNAi library for genes that, when knocked down by RNAi, changed the expression of SOD1.  One prominent hit was TDP-43, which, when reduced, led to a three-fold increase in expression of SOD1.

The increase in SOD1 by TDP-43 knockdown “could result because of alterations in transcription or mRNA stability brought about by TDP-43,” the authors stated.  “However the effect could also be mediated indirectly through an unknown factor which may or may not involve splicing, another known function of TDP-43.”  The genes and proteins in this study were the wild-type, non-mutated forms.  Future work will be needed to determine whether mutation of one or the other gene alters the functional effect identified here.

Visit http://www.ncbi.nlm.nih.gov/pubmed/22563406 for more information on “TDP-43 Identified from a Genome Wide RNAi Screen for SOD1 Regulators” PLoS One. 2012;7(4):e35818. Epub 2012 Apr 26 by B.R. Somalinga, C.E. Day, S. Wei, M.G. Roth and P.J. Thomas.

New TDP-43 Mouse Model Develops Inclusions, Neurodegeneration from Late Expression
The timing of TDP-43 expression plays a critical role in its effect on brain development and neurodegeneration, according to new research funded by The ALS Association.  The results are important both for understanding the effects of this ALS-causing gene and for development of mouse models that accurately mimic the human disease.

The authors developed a conditional-expression mouse model, in which timing of expression of wild-type human TDP-43 in the forebrain was controlled by a genetic switch built into the gene.  Continuous expression during early neuronal development resulted in a severe and rapidly progressing phenotype and early lethality, suggesting that “neurons in the developing forebrain are extremely sensitive to TDP-43 overexpression and that timing of TDP-43 overexpression in transgenic mice must be considered when distinguishing normal roles of TDP-43,” they wrote.

In contrast, overexpression in adulthood produced a slowly progressive neurodegeneration, progressive gliosis, and cytoplasmic inclusions similar to those seen in ALS and frontotemporal dementia in which TDP-43 plays a role.

“The availability of this new TDP-43 model system provides the field with a more pathologically similar transgenic mouse model for [TDP-related frontotemporal dementia] as well as a system in which the role of TDP-43 in development versus disease can now be distinguished,” they concluded.

Visit http://www.ncbi.nlm.nih.gov/pubmed/22539017 for more information on “Neuronal sensitivity to TDP-43 overexpression is dependent on timing of induction” Acta Neuropathol. 2012 Jun;123(6):807-23. Epub 2012 Apr 27 by A. Cannon, B. Yang, J. Knight, I.M Farnham, Y. Zhang, C.A. Wuertzer, S. D'Alton, W.L. Lin, M. Castanedes-Casey, L. Rousseau, B. Scott, M. Jurasic, J. Howard, X. Yu, R. Bailey, M.R. Sarkisian, D.W. Dickson, L Petrucelli, and J. Lewis.

Visualizing Corticospinal Motor Neurons Reveals Early Degeneration
Corticospinal motor neurons (CSMNs) can be selectively targeted for gene delivery, according to new research supported by The ALS Association.  The development of this selective targeting should allow researchers to better characterize the disease process in these neurons and potentially to deliver therapy to them.  CSMNs send movement instructions from the brain to the spinal cord.

Researchers delivered adeno-associated virus type 2 (AAV2) to the corticospinal tract in both wild-type and SOD1-G93A mice.  The virus was selectively taken up by CSMNs, and retrogradely transported to the cortex, allowing visualization of motor cortical neurons in both health and disease.  The authors discovered that CSMNs displayed “severe and selective apical dendrite degeneration in SOD1 mice,” possibly indicating the importance of this process as an early stage in ALS.  The ability to label CSMNs and transfect them with the gene-carrying AAVs may lead to better understanding of the early steps of ALS in the brain.

Visit http://www.ncbi.nlm.nih.gov/pubmed/22521461 for more information about “ AAV2 mediated retrograde transduction of corticospinal motor neurons reveals initial and selective apical dendrite degeneration in ALS” Neurobiol Dis. 2012 Apr 11 by J.H. Jara, S.R. Villa, N.A. Khan, M.C. Bohn and P.H. Ozdinler.

Angiogenin from Motor Neurons Is Taken Up by Astrocytes, Cleaves RNA
Motor neurons produce and release angiogenin, which is subsequently taken up by neighboring astrocytes where it cleaves small RNAs, but that cleavage is reduced by ALS-associated angiogenin mutants, according to new research funded by The ALS Association.  The results provide further evidence of the importance of astrocyte-motor neuron interaction in ALS.

Angiogenin is best known as a factor promoting blood vessel growth, but it has numerous other functions as well, most or all of which arise from its basic activity as an enzyme that cleaves RNA.  In 2006, researchers found that some cases of both familial and sporadic ALS were linked to mutations in the angiogenin gene.

To better understand the potential molecular mechanisms involved in angiogenin’s effect, researchers studied its trafficking between motor neurons and astrocytes in cell culture.  They found that neurons secreted angiogenin, which was taken up by astrocytes, where it cleaved small RNAs within the astrocytes.  Cleavage, and to a lesser extent uptake, was reduced by ALS-associated mutations.

“Our results…suggest that angiogenin exerts important additional biological functions by acting in paracrine (from one cell to a neighboring one), and that angiogenin regulates astroglia function by targeting novel subsets of RNAs,” the authors concluded.  “Future studies will need to identify and characterize these novel angiogenin RNA substrates.”  Small RNAs are increasingly being recognized as important regulators of many cellular functions, especially gene expression, suggesting that angiogenin may mediate influence one or more pathways in astrocytes through its ability to cleave these small regulatory RNAs.

Visit http://www.ncbi.nlm.nih.gov/pubmed/22496549 for more information about “Motoneurons secrete angiogenin to induce RNA cleavage in astroglia” J Neurosci. 2012 Apr 11;32(15):5024-38 by A. Skorupa, M.A. King, I.M. Aparicio, H. Dussmann, K. Coughlan, B. Breen, D. Kieran, C.G Concannon, P. Marin and J.H. Prehn.

Testosterone Derivative Improves Muscle Bulk and Strength in ALS Mice
A testosterone-like molecule increases muscle bulk and strength in SOD1 mice when administered shortly after the onset of symptoms, according to research funded by The ALS Association.  Testosterone is the male sex hormone and is a type of anabolic steroid, whose muscle-building effects are well known.  “Considering that muscle-related symptoms are closely related with the quality of the patient’s life, targeting ALS-affected muscle may provide practical benefits to patients,” the authors hypothesized.

As an initial test of that hypothesis, they implanted a silicone tube containing 5-alpha-dihydrotestosterone (DHT), a breakdown product of testosterone in SOD1 mouse at the early symptomatic stage, 75 days after birth.  Only male mice were treated.  Compared to untreated SOD1 mice, treatment increased the weight of leg muscles by 30-40%, increased the muscle cross-sectional area by about 20%, and increased grip strength by up to 40%.  Body weight was also moderately increased.  The authors also showed that treatment increased the expression of insulin-like growth factor 1 and 2, which are known to mediate increases in muscle bulk.  Treated mice performed better on a test of motor performance and lived about 5% longer than untreated mice.

“The beneficial effect of DHT illustrated in the present study may provide an initial step toward a possible therapy for ALS,” the authors concluded.  “However, several concerns remain regarding adverse side effects of chronic DHT treatment,” including increased risk of prostate cancer and the effects of DHT in women.  Nonetheless, they argue, the potential of DHT or related steroids to benefit ALS patients warrant further investigation.

Visit http://www.plosone.org/article/info%3Adoi%2F10.1371%2Fjournal.pone.0037258 and http://www.ncbi.nlm.nih.gov/pubmed/22606355 for more information about “Dihydrotestosterone ameliorates degeneration in muscle, axons and motoneurons and improves motor function in amyotrophic lateral sclerosis model mice” PLoS One. 2012;7(5):e37258. Epub 2012 May 14 by Y.E. Yoo and C.P. Ko.