Cindy Chen

Alzheimer's disease (AD) and familial Danish dementia (FDD) are degenerative neurological diseases characterized by amyloid pathology. Normal human sera contain IgG antibodies that specifically bind diverse preamyloid and amyloid proteins and have shown therapeutic potentialin vitroandin vivo. We cloned one of these antibodies, 3H3, from memory B cells of a healthy individual using a hybridoma method. 3H3 is an affinity-matured IgG that binds a pan-amyloid epitope, recognizing both Aβ and λ Ig light chain (LC) amyloids, which are associated with AD and primary amyloidosis, respectively. The pan-amyloid-binding properties of 3H3 were demonstrated using ELISA, immunohistochemical studies, and competition binding assays. Functional studies showed that 3H3 inhibits both Aβ and LC amyloid formationin vitroand abrogates disruption of hippocampal synaptic plasticity by AD-patient-derived soluble Aβin vivo. A 3H3 single-chain variable fragment (scFv) retained the binding specificity of the 3H3 IgG and, when expressed in the brains of transgenic mice using an adeno-associated virus (AAV) vector, decreased parenchymal Aβ amyloid deposition in TgCRND8 mice and ADan (Danish Amyloid) cerebral amyloid angiopathy in the mouse model of FDD. These data indicate that naturally occurring human IgGs can recognize a conformational, amyloid-specific epitope and have potent anti-amyloid activities, providing a rationale to test their potential as antibody therapeutics for diverse neurological and other amyloid diseases.

The anti‐inflammatory and neuroprotective effects of estrogen on multiple sclerosis (MS) have been reported in previous studies. Evidence has been found that estrogen can inhibit axonal loss in the MOG‐induced experimental autoimmune encephalomyelitis (EAE) model of MS. Rho‐kinase (ROCK) mediates axonal growth‐inhibitory signals via the Rho/Rho‐kinase pathway. The inhibition of ROCK decreases axonal loss in EAE. However, there is no study reporting the association between estrogen and ROCK in MS and EAE. We examined the anti‐inflammatory and axonal protective effects of estrogen and explored the probable mechanism whereby estrogen inhibits ROCK through ERα in EAE rats. Results show that 17β‐estradiol (E2) can significantly avoid the loss of neurological function in EAE rats. E2 also decreased the infiltration of inflammatory cells and cytokines including IL‐1β, TNF‐α, and IL‐17, but increased the expression of IL‐4. E2 inhibited the expression of ROCK and NF‐200 in EAE rats. The inhibitory effect on ROCK was abolished when nonselective estrogen receptor (ER) antagonist ICI 182780 was added to E2. Furthermore, the expression of ROCK was inhibited by ERα‐selective ligand agonist propyl pyrazole triol. ERβ‐selective ligand WAY‐202041 has no effect on the expression of ROCK. These observations suggest that estrogen inhibits the expression of ROCK in EAE rats and that the inhibitory effects are mediated by ERα rather than ERβ. Synapse 67:399–406, 2013.

Minocycline has been recently implicated in protection against focal cerebral ischemia reperfusion (I/R), but the protective effects on neurobehavioral abnormalities remains contradictory. In the present study, we investigate whether minocycline improves axonal regeneration and neurological function recovery by inhibiting the expression of the repulsive guidance molecular A (RGMa) after focal cerebral ischemia reperfusion. Male Sprague‐Dawley (SD) rats were subjected to occlusion of the right middle cerebral artery (MCAO) for 2 h and 3 mg kg⁻¹ minocycline was injected intravenously immediately after reperfusion twice a day for 14 days. The staircase test and modified neurological severity score (mNSS) were performed to evaluate functional outcome and blood‐brain barrier (BBB) permeability was assessed by Evan's blue dye extravasation (EB) at the expected time point. The expression of RGMa in ischemic cortex was measured by immunohistochemical staining and Western blot 2 weeks after MCAO. Neurofilament protein 200 (NF‐200) immunohistochemical staining was used to assess axonal damage. Treatment with minocycline at a dose of 3 mg kg⁻¹ via the caudal vein significantly reduced the extravasation of EB, elevated mNSS and improved forelimb motor function as assessed by the staircase test when compared to the I/R group (P < 0.05). Moreover, axonal regrowth was enhanced in the minocycline treatment group when compared to the I/R group (P < 0.05). In addition, minocycline significantly reduced the expression of RGMa protein 2 weeks after MCAO as assessed by both immunostaining and Western blot. Our studies suggest that early minocycline treatment promotes neurological functional recovery and axonal regeneration in rats after MCAO, which might be mediated by down‐regulating RGMa expression. Synapse, 2013.

Botulinum neurotoxin (BoNT) potently inhibits cholinergic signaling at the neuromuscular junction. The ideal countermeasures for BoNT exposure are monoclonal antibodies or BoNT antisera, which form BoNT-containing immune complexes that are rapidly cleared from the general circulation. Clearance of opsonized toxins may involve complement receptor-mediated immunoadherence to red blood cells (RBC) in primates or to platelets in rodents. Methods of enhancing immunoadherence of BoNT-specific antibodies may increase their potency in vivo. We designed a novel fusion protein (FP) to link biotinylated molecules to glycophorin A (GPA) on the RBC surface. The FP consists of an scFv specific for murine GPA fused to streptavidin. FP:mAb:BoNT complexes bound specifically to the RBC surface in vitro. In a mouse model of BoNT neutralization, the FP increased the potency of single and double antibody combinations in BoNT neutralization. A combination of two antibodies with the FP gave complete neutralization of 5,000 LD50 BoNT in mice. Neutralization in vivo was dependent on biotinylation of both antibodies and correlated with a reduction of plasma BoNT levels. In a post-exposure model of intoxication, FP:mAb complexes gave complete protection from a lethal BoNT/A1 dose when administered within 2 hours of toxin exposure. In a pre-exposure prophylaxis model, mice were fully protected for 72 hours following administration of the FP:mAb complex. These results demonstrate that RBC-targeted immunoadherence through the FP is a potent enhancer of BoNT neutralization by antibodies in vivo.