CD34 Identifies a Subset of Proliferating Microglial Cells Associated with Degenerating Motor Neurons in ALS

Abstract

Amyotrophic lateral sclerosis (ALS) is characterized by degeneration of upper and lower motor neurons accompanied by proliferation of reactive microglia in affected regions. However, it is unknown whether the hematopoietic marker CD34 can identify a subpopulation of proliferating microglial cells in the ALS degenerating spinal cord. Immunohistochemistry for CD34 and microglia markers was performed in lumbar spinal cords of ALS rats bearing the SOD1G93A mutation and autopsied ALS and control human subjects. Characterization of CD34-positive cells was also performed in primary cell cultures of the rat spinal cords. CD34 was expressed in a large number of cells that closely interacted with degenerating lumbar spinal cord motor neurons in symptomatic SOD1G93A rats, but not in controls. Most CD34+ cells co-expressed the myeloid marker CD11b, while only a subpopulation was stained for Iba1 or CD68. Notably, CD34+ cells actively proliferated and formed clusters adjacent to damaged motor neurons bearing misfolded SOD1. CD34+ cells were identified in the proximity of motor neurons in autopsied spinal cord from sporadic ALS subjects but not in controls. Cell culture of symptomatic SOD1G93A rat spinal cords yielded a large number of CD34+ cells exclusively in the non-adherent phase, which generated microglia after successive passaging. A yet unrecognized CD34+ cells, expressing or not the microglial marker Iba1, proliferate and accumulate adjacent to degenerating spinal motor neurons, representing an intriguing cell target for approaching ALS pathogenesis and therapeutics.

‘A picture is worth a thousand words’: The use of microscopy for imaging neuroinflammation

“…CD45 could help in this case as it is more expressed in monocytes than in microglia, so both Iba-1-and CD45-positive cells -particularly round-shaped cells -are probably monocytes, while Iba-1-positive and CD45-negative cells with processes are microglia [62,64]. Another helpful marker is CD68 (a macrophage/phagocytic activity marker) which appears after ischemia in the border zone of the lesion, but invades the ischemia core 7 days after the insult [56,62].…”

“…Usually, microglia alterations are widely used to suggest a neuroinflammation condition in the CNS using the ionized calcium-binding adaptor molecule 1 (Iba-1) marker [54][55][56]. Some authors applied double-labeling of the Iba-1 as a marker for all microglial cells, labeling the soma, with LN3 (a monoclonal antibody used as a microglial activation marker as it recognizes the major histocompatibility complex (MHC)-II), to evaluate microglial hypertrophic and ameboid morphology [57].…”

“…Although it can lead to confusion with neutrophils, CD11b is used as a marker for microglia, as these cells express this protein [56,62]. Conversely, myeloperoxidase protein (MPO) can be used as a marker for neutrophils [63], allowing discrimination of neutrophils from microglia.…”

A Nitroalkene Benzoic Acid Derivative Targets Reactive Microglia and Prolongs Survival in an Inherited Model of ALS via NF-κB Inhibition

“…Human Spinal Cord Immunohistochemistry As previously described [34], 10-μm spinal cord paraffin sections were sliced using a microtome. Following deparaffinization, slices were blocked and permeabilized in BSA 5% (v/v)/Triton X-100 0.5% (v/v) for 2 h at room temperature.…”

Review: Microglia in motor neuron disease

“…A population of microglia, which are immunopositive for the haemopoetic transmembrane phosphoglycoprotein, CD34, have been observed in the ventral horn of sMND spinal cord [56]. In control cases, immunoreactivity for CD34 is restricted to the blood vessels and capillaries.…”