Monoclonal enolase-1 blocking antibody ameliorates pulmonary inflammation and fibrosis.

BACKGROUND: Idiopathic pulmonary fibrosis (IPF) is a chronic fatal disease with limited therapeutic options. The infiltration of monocytes and fibroblasts into the injured lungs is implicated in IPF. Enolase-1 (ENO1) is a cytosolic glycolytic enzyme which could translocate onto the cell surface and act as a plasminogen receptor to facilitate cell migration via plasmin activation. Our proprietary ENO1 antibody, HL217, was screened for its specific binding to ENO1 and significant inhibition of cell migration and plasmin activation (patent: US9382331B2). METHODS: In this study, effects of HL217 were evaluated in vivo and in vitro for treating lung fibrosis. RESULTS: Elevated ENO1 expression was found in fibrotic lungs in human and in bleomycin-treated mice. In the mouse model, HL217 reduced bleomycin-induced lung fibrosis, inflammation, body weight loss, lung weight gain, TGF-ß upregulation in bronchial alveolar lavage fluid (BALF), and collagen deposition in lung. Moreover, HL217 reduced the migration of peripheral blood mononuclear cells (PBMC) and the recruitment of myeloid cells into the lungs. In vitro, HL217 significantly reduced cell-associated plasmin activation and cytokines secretion from primary human PBMC and endothelial cells. In primary human lung fibroblasts, HL217 also reduced cell migration and collagen secretion. CONCLUSIONS: These findings suggest multi-faceted roles of cell surface ENO1 and a potential therapeutic approach for pulmonary fibrosis.

High-Frequency Stimulation of the Subthalamic Nucleus Activates Motor Cortex Pyramidal Tract Neurons by a Process Involving Local Glutamate, GABA and Dopamine Receptors in Hemi-Parkinsonian Rats.

Deep brain stimulation (DBS) is widely used to treat advanced Parkinson's disease (PD). Here, we investigated how DBS applied on the subthalamic nucleus (STN) influenced the neural activity in the motor cortex. Rats, which had the midbrain dopaminergic neurons partially depleted unilaterally, called the hemi-Parkinsonian rats, were used as a study model. c-Fos expression in the neurons was used as an indicator of neural activity. Application of high-frequency stimulation (HFS) upon the STN was used to mimic the DBS treatment. The motor cortices in the two hemispheres of hemi-Parkinsonian rats were found to contain unequal densities of c-Fos-positive (Fos+) cells, and STN-HFS rectified this bilateral imbalance. In addition, STN-HFS led to the intense c-Fos expression in a group of motor cortical neurons which exhibited biochemical and anatomical characteristics resembling those of the pyramidal tract (PT) neurons sending efferent projections to the STN. The number of PT neurons expressing high levels of c-Fos was significantly reduced by local application of the antagonists of non-N-methyl-D-aspartate (non-NMDA) glutamate receptors, gammaaminobutyric acid A (GABAA) receptors and dopamine receptors in the upper layers of the motor cortex. The results indicate that the coincident activations of synapses and dopamine receptors in the motor cortex during STN-HFS trigger the intense expression of c-Fos of the PT neurons. The implications of the results on the cellular mechanism underlying the therapeutic effects of STN-DBS on the movement disorders of PD are also discussed.

A Novel Enolase-1 Antibody Targets Multiple Interacting Players in the Tumor Microenvironment of Advanced Prostate Cancer.

Prostate cancer is one of the most common causes of cancer death in men worldwide, and the treatment options are limited for patients with advanced stages of prostate cancer. Upon oncogenic or inflammatory stimulation, tumor cells or immune cells express cell surface enolase-1 (ENO1) as plasminogen receptor to facilitate their migration via plasmin activation. Little is known about the roles of ENO1 in prostate cancer, especially in the tumor microenvironment (TME). We hypothesized that targeting surface ENO1 with specific mAbs would exert multifactorial therapeutic potentials against prostate cancer. In vivo, we showed ENO1 mAb (HuL227) reduced the growth of subcutaneous PC-3 xenograft, monocytes recruitment, and intratumoral angiogenesis. In a PC-3 intratibial implantation model, HuL227 reduced tumor growth and osteoclast activation in the bone. To investigate the antitumor mechanism of ENO1 mAb, we found that blocking surface ENO1 significantly reduced VEGF-A-induced tube formation of endothelial cells in vitro. Furthermore, HuL227 inhibited inflammation-enhanced osteoclasts activity and the secretion of invasion-related cytokines CCL2 and TGFß from osteoclasts. In addition, inflammation-induced migration and chemotaxis of androgen-independent prostate cancer cells were dose-dependently inhibited by HuL227. In summary, we showed that, ENO1 mAb targets multiple TME niches involved in prostate cancer progression and bone metastasis via a plasmin-related mechanism, which may provide a novel immunotherapy approach for men with advanced prostate cancer.

Haloperidol Instigates Endometrial Carcinogenesis and Cancer Progression by the NF-κB/CSF-1 Signaling Cascade.

Haloperidol is a routine drug for schizophrenia and palliative care of cancer; it also has antitumor effects in several types of cancer. However, the role of haloperidol in endometrial cancer (EC) development is still unclear. Here, we show that chronic haloperidol treatment in clinically relevant doses induced endometrial hyperplasia in normal mice and promoted tumor growth and malignancy in mice with orthotopic EC. The pharmacokinetic study indicated that haloperidol highly accumulated in the uterus of mice. In vitro studies revealed that haloperidol stimulated the cellular transformation of human endometrial epithelial cells (HECCs) and promoted the proliferation, migration, and invasion of human endometrial carcinoma cells (HECCs) by activating nuclear factor kappa B (NF-κB) and its downstream signaling target, colony-stimulating factor 1 (CSF-1). Gain of function of CSF-1 promotes the cellular transformation of HEECs and the malignant progression of HECCs. Moreover, blockade of CSF-1 inhibited haloperidol-promoted EC progression in vitro and in vivo. A population-based cohort study of EC patients further demonstrated that the use of haloperidol was associated with increased EC-specific mortality. Collectively, these findings indicate that clinical use of haloperidol could potentially be harmful to female patients with EC.

Muscle Tissue Damage and Recovery After EV71 Infection Correspond to Dynamic Macrophage Phenotypes.

Enterovirus 71 (EV71) is a positive single-stranded RNA virus from the enterovirus genus of the Picornaviridae family. Most young children infected with EV71 develop mild symptoms of hand, foot and mouth disease, but some develop severe symptoms with neurological involvement. Limb paralysis from EV71 infection is presumed to arise mainly from dysfunction of motor neurons in the spinal cord. However, EV71 also targets and damages skeletal muscle, which may also contribute to the debilitating symptoms. In this study, we have delineated the impacts of EV71 infection on skeletal muscle using a mouse model. Mouse pups infected with EV71 developed limb paralysis, starting at day 3 post-infection and peaking at day 5-7 post-infection. At later times, mice recovered gradually but not completely. Notably, severe disease was associated with high levels of myositis accompanied by muscle calcification and persistent motor end plate abnormalities. Interestingly, macrophages exhibited a dynamic change in phenotype, with inflammatory macrophages (CD45+CD11b+Ly6Chi) appearing in the early stage of infection and anti-inflammatory/restorative macrophages (CD45+CD11b+Ly6Clow/-) appearing in the late stage. The presence of inflammatory macrophages was associated with severe inflammation, while the restorative macrophages were associated with recovery. Altogether, we have demonstrated that EV71 infection causes myositis, muscle calcification and structural defects in motor end plates. Subsequent muscle regeneration is associated with a dynamic change in macrophage phenotype.

Quantitative evaluation of motor function before and after engraftment of dopaminergic neurons in a rat model of Parkinson's disease.

Although gait change is considered a useful indicator of severity in animal models of Parkinson's disease, systematic and extensive gait analysis in animal models of neurological deficits is not well established. The CatWalk-assisted automated gait analysis system provides a comprehensive way to assess a number of dynamic and static gait parameters simultaneously. In this study, we used the Catwalk system to investigate changes in gait parameters in adult rats with unilateral 6-OHDA-induced lesions and the rescue effect of dopaminergic neuron transplantation on gait function. Four weeks after 6-OHDA injection, the intensity and maximal area of contact were significantly decreased in the affected paws and the swing speed significantly decreased in all four paws. The relative distance between the hind paws also increased, suggesting that animals with unilateral 6-OHDA-induced lesions required all four paws to compensate for loss of balance function. At 8 weeks post-transplantation, engrafted dopaminergic neurons expressed tyrosine hydroxylase. In addition, the intensity, contact area, and swing speed of the four limbs increased and the distance between the hind paws decreased. Partial recovery of methamphetamine-induced rotational response was also noted.

A Computationally-Efficient, Online-Learning Algorithm for Detecting High-Voltage Spindles in the Parkinsonian Rats.

Abnormally-synchronized, high-voltage spindles (HVSs) are associated with motor deficits in 6-hydroxydopamine-lesioned parkinsonian rats. The non-stationary, spike-and-wave HVSs (5-13 Hz) represent the cardinal parkinsonian state in the local field potentials (LFPs). Although deep brain stimulation (DBS) is an effective treatment for the Parkinson's disease, continuous stimulation results in cognitive and neuropsychiatric side effects. Therefore, an adaptive stimulator able to stimulate the brain only upon the occurrence of HVSs is demanded. This paper proposes an algorithm not only able to detect the HVSs with low latency but also friendly for hardware realization of an adaptive stimulator. The algorithm is based on autoregressive modeling at interval, whose parameters are learnt online by an adaptive Kalman filter. In the LFPs containing 1131 HVS episodes from different brain regions of four parkinsonian rats, the algorithm detects all HVSs with 100% sensitivity. The algorithm also achieves higher precision (96%) and lower latency (61 ms), while requiring less computation time than the continuous wavelet transform method. As the latency is much shorter than the mean duration of an HVS episode (4.3 s), the proposed algorithm is suitable for realization of a smart neuromodulator for mitigating HVSs effectively by closed-loop DBS.

TLR7 Is Critical for Anti-Viral Humoral Immunity to EV71 Infection in the Spinal Cord.

Enterovirus 71 (EV71) is a positive single-stranded RNA (ssRNA) virus from the enterovirus genus of Picornaviridae family and causes diseases ranged from the mild disease of hand, foot and mouth disease (HFMD) to the severe disease of neurological involvement in young children. TLR7 is an intracellular pattern recognition receptor (PRR) recognizing viral ssRNA. In this study, we investigated the role of TLR7 in EV71 infection in mouse pups (10-12 days old) and found that wild-type (WT) and TLR7 knock-out (TLR7KO) mice infected with EV71 showed similar limb paralysis at the onset and peak of the disease, comparable loss of motor neurons, and similar levels of antiviral molecules in the spinal cord. These results suggest that TLR7 is not the absolute PRR for EV71 in the spinal cord. Interestingly, TLR7KO mice infected with EV71 exhibited significantly delayed recovery from limb paralysis compared with WT mice. TLR7KO mice infected with EV71 showed significantly decreased levels of IgM and IgG2, important antibodies for antiviral humoral immunity. Furthermore, TLR7KO mice infected with EV71 showed a decrease of germinal center B cells in the spleen compared with WT mice. Altogether, our study suggests that TLR7 plays a critical role in anti-viral humoral immunity rather than in being a PRR in the spinal cord during EV71 infection in young mice.

A Battery-Less, Implantable Neuro-Electronic Interface for Studying the Mechanisms of Deep Brain Stimulation in Rat Models.

Although deep brain stimulation (DBS) has been a promising alternative for treating several neural disorders, the mechanisms underlying the DBS remain not fully understood. As rat models provide the advantage of recording and stimulating different disease-related regions simultaneously, this paper proposes a battery-less, implantable neuro-electronic interface suitable for studying DBS mechanisms with a freely-moving rat. The neuro-electronic interface mainly consists of a microsystem able to interact with eight different brain regions bi-directionally and simultaneously. To minimize the size of the implant, the microsystem receives power and transmits data through a single coil. In addition, particular attention is paid to the capability of recording neural activities right after each stimulation, so as to acquire information on how stimulations modulate neural activities. The microsystem has been fabricated with the standard 0.18 µm CMOS technology. The chip area is 7.74 mm (2) , and the microsystem is able to operate with a single supply voltage of 1 V. The wireless interface allows a maximum power of 10 mW to be transmitted together with either uplink or downlink data at a rate of 2 Mbps or 100 kbps, respectively. The input referred noise of recording amplifiers is 1.16 µVrms, and the stimulation voltage is tunable from 1.5 V to 4.5 V with 5-bit resolution. After the electrical functionality of the microsystem is tested, the capability of the microsystem to interface with rat brain is further examined and compared with conventional instruments. All experimental results are presented and discussed in this paper.