Persistent Malfunction of Glymphatic and Meningeal Lymphatic Drainage in a Mouse Model of Subarachnoid Hemorrhage.

Subarachnoid hemorrhage (SAH) is a devastating cerebrovascular event that often is followed by permanent brain impairments. It is necessary to explore the pathogenesis of secondary pathological damages in order to find effective interventions for improving the prognosis of SAH. Blockage of brain lymphatic drainage has been shown to worsen cerebral ischemia and edema after acute SAH. However, whether or not there is persistent dysfunction of cerebral lymphatic drainage following SAH remains unclear. In this study, autologous blood was injected into the cisterna magna of mice to establish SAH model. One week after surgery, SAH mice showed decreases in fluorescent tracer drainage to the deep cervical lymph nodes (dcLNs) and influx into the brain parenchyma after injection into the cisterna magna. Moreover, SAH impaired polarization of astrocyte aquaporin-4 (AQP4) that is a functional marker of glymphatic clearance and resulted in accumulations of Tau proteins as well as CD3+, CD4+, and CD8+ cells in the brain. In addition, pathological changes, including microvascular spasm, activation of glial cells, neuroinflammation, and neuronal apoptosis were observed in the hippocampus of SAH mice. Present results demonstrate persistent malfunction of glymphatic and meningeal lymphatic drainage and related neuropathological damages after SAH. Targeting improvement of brain lymphatic clearance potentially serves as a new strategy for the treatment of SAH.

Blocking meningeal lymphatic drainage aggravates Parkinson's disease-like pathology in mice overexpressing mutated α-synuclein.

BACKGROUND: Abnormal aggregation of brain α-synuclein is a central step in the pathogenesis of Parkinson's disease (PD), thus, it is reliable to promote the clearance of α-synuclein to prevent and treat PD. Recent studies have revealed an essential role of glymphatic system and meningeal lymphatic vessels in the clearance of brain macromolecules, however, their pathophysiological aspects remain elusive. METHOD: Meningeal lymphatic drainage of 18-week-old A53T mice was blocked via ligating the deep cervical lymph nodes. Six weeks later, glymphatic functions and PD-like phenotypes were systemically analyzed. RESULTS: Glymphatic influx of cerebrospinal fluid tracer was reduced in A53T mice, accompanied with perivascular aggregation of α-synuclein and impaired polarization of aquaporin 4 expression in substantia nigra. Cervical lymphatic ligation aggravated glymphatic dysfunction of A53T mice, causing more severe accumulation of α-synuclein, glial activation, inflammation, dopaminergic neuronal loss and motor deficits. CONCLUSION: The results suggest that brain lymphatic clearance dysfunction may be an aggravating factor in PD pathology.

Aquaporin-4-dependent glymphatic solute transport in the rodent brain.

The glymphatic system is a brain-wide clearance pathway; its impairment contributes to the accumulation of amyloid-ß. Influx of cerebrospinal fluid (CSF) depends upon the expression and perivascular localization of the astroglial water channel aquaporin-4 (AQP4). Prompted by a recent failure to find an effect of Aqp4 knock-out (KO) on CSF and interstitial fluid (ISF) tracer transport, five groups re-examined the importance of AQP4 in glymphatic transport. We concur that CSF influx is higher in wild-type mice than in four different Aqp4 KO lines and in one line that lacks perivascular AQP4 (Snta1 KO). Meta-analysis of all studies demonstrated a significant decrease in tracer transport in KO mice and rats compared to controls. Meta-regression indicated that anesthesia, age, and tracer delivery explain the opposing results. We also report that intrastriatal injections suppress glymphatic function. This validates the role of AQP4 and shows that glymphatic studies must avoid the use of invasive procedures.

Ginkgolide K promotes angiogenesis in a middle cerebral artery occlusion mouse model via activating JAK2/STAT3 pathway.

Ginkgolide K (GK) is a new compound extracted from the leaves of Ginkgo biloba, which has been recognized to exert anti-oxidative stress and neuroprotective effect on ischemic stroke. While whether it plays an enhanced effect on angiogenesis during ischemic stroke remains unknown. The aim of this study was to investigate the effect of ginkgolide K on promoting angiogenesis as well as the protective mechanism after cerebral ischemia-reperfusion. Using the transient middle cerebral artery occlusion (tMCAO) mouse model, we found that GK (3.5, 7.0, 14.0 mg/kg, i.p., bid., 2 weeks) attenuated neurological impairments, and promoted angiogenesis of injured ipsilateral cortex and striatum after 14 days of cerebral ischemia-reperfusion in mice. Further, GK (3.5 mg/kg in vivo, 10 µM in vitro) significantly up-regulated the expressions of HIF-1α and VEGF in tMCAO mouse brains and in b End3 cells after OGD/R, and GK-induced upregulation of HIF-1α and VEGF in b End3 cells could be abolished by JAK2/STAT3 inhibitor AG490. Our results demonstrate that GK promotes angiogenesis after ischemia stroke through increasing the expression of HIF-1α/VEGF via JAK2/STAT3 pathway, which provide an insight into the novel clinical application of GK and its analogs in ischemic stroke therapy in future.

Expression, purification, and characterization of recombinant human H-chain ferritin.

Based on their nanocage architectures, ferritins show their potential applications in medical imaging and therapeutic delivery systems. However, the recombinant human H-chain ferritin (rHF) is prone to form inclusion bodies in Escherichia coli. In our study, the cDNA of rHF was cloned into plasmid pET28a under the control of a T7 promoter. Molecular chaperones, including GroES, GroEL, and trigger factor, were coexpressed with rHF to facilitate its correct folding. The results showed that the solubility of rHF was increased more than threefold with the help of molecular chaperones. Taking advantages of its N-terminal His-tag, rHF was then purified with Ni-affinity chromatography. With a yield of 15 mg/L from bacterial culture, the purified rHF was analyzed by circular dichroism spectrometry for its secondary structure. Moreover, the rHF nanocages were characterized by transmission electron microscopy and dynamic light scattering. Our results indicate that rHF is able to self-assemble into nanocages with a narrow size distribution.

Expression, purification, and characterization of recombinant human L-chain ferritin.

Ferritins form nanocage architectures and demonstrate their potential to serve as functional nanomaterials with potential applications in medical imaging and therapy. In our study, the cDNA of human L-chain ferritin was cloned into plasmid pET-28a for its overexpression in Escherichia coli. However, the recombinant human L-chain ferritin (rLF) was prone to form inclusion bodies. Molecular chaperones were co-expressed with rLF to facilitate its correct folding. Our results showed that the solubility of rLF was increased about 3-fold in the presence of molecular chaperones, including GroEL, GroES and trigger factor. Taking advantage of its N-terminal His-tag, rLF was then purified with Ni-affinity chromatography. With a yield of 10 mg/L from bacterial culture, the purified rLF was analyzed by circular dichroism spectrometry for its secondary structure. Furthermore, the rLF nanocages were characterized using dynamic light scattering and transmission electron microscopy.

Fused hydrophobic elastin-like-peptides (ELP) enhance biological activity of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL).

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a promising anticancer agent with tumor-selective apoptotic activity. Formation of aggregates as trimer is the prerequisite for TRAIL's function as an apoptosis inducer. However the polymerization property of TRAIL has also brought difficulties for its production. RGD-TRAIL is an integrin-targeting TRAIL mutant with enhanced apoptosisinducing activity towards tumor cells both in vitro and in vivo. When expressed in E. coli, TRAIL or its mutant RGDTRAIL usually formed inclusion bodies. Their extreme aggregation propensity for aggregation destabilizes the protein, leading to poor recovery and therefore low yield from the purification process. The low purification efficiency of TRAIL retards its industrial application and large-scale production. To avoid the above problems during RGD-TRAIL production, we employed elastin-like polypeptides (ELPs) for the fusion-expression of recombinant RGD-TRAIL. Recombinant RGD-TRAIL-ELP was expressed in a soluble form and efficiently purified from the clarified cell extracts by three rounds of inverse transition cycling (ITC). SDS-PAGE and Western blotting analyses of purified RGD-TRAIL-ELP showed that RGD-TRAIL-ELP was successfully purified and the yield was up to 10 mg/L of bacterial culture. Apoptosis assay was performed in human colorectal carcinoma cells (COLO-205) and human breast cancer cell line (MDA-MB-231) to assess the potency of the fusion protein. Fusion with hydrophobic ELP effectively enhanced RGD-TRAIL's biological activity. The higher activity and appropriate particle size of RGD-TRAIL-ELP could be used for RGD-TRAIL delivery in tumor therapy.

[Clinical analyses of risk factors related to ectopic pregnancy].

OBJECTIVE: To conduct an epidemiological survey on the relevant risk factors of ectopic pregnancy (EP) and provide scientific rationales for prevention and reducing its incidence. METHODS: During June 2010 to December 2011 at three local hospitals, a total of 800 patients with a diagnosis of EP responded to a questionnaire survey for understanding the risk factors of EP. And another 700 cases of normal early pregnancy women were selected as control group. Both multi and uni-factorial regression analyses were performed for the acquired clinical data. And the high-risk factors of EP were screened. RESULTS: Among them, the age distribution was ≤ 25 years (n = 175, 25.5%), 26-30 years (n = 302, 37.8%) and 31-35 years (n = 213, 26.6%). No contraceptive measure was adopted for 259 women (32.4%). The risk factors related to EP included oral emergency contraceptives, intrauterine device (IUD), pelvic inflammatory disease, infertility history, previous EP, smoking and age, etc. Based upon multivariate Logistic regression analysis of screening results, the decreasing odds rations were infertility, EP history, smoking history, emergency contraceptive use, history of EP, IUD, pelvic inflammatory disease and age. CONCLUSION: The risks of EP are affected by many factors, including infertility, EP history, smoking history, emergency contraceptive use, history of EP, IUD, pelvic inflammatory disease and age.

[Research on the feasibility of a magnetic-coupling-driven axial flow blood pump].

A new-designed axial flow blood pump, dived by magnetic coupling and using internal hollow brushless DC motor and inlet and outlet in line with impeller, was tested in mimic circuit. The results showed good performance of the new pump and indicated that its hydrodynamic characteristic can meet the demands of clinical extracorporeal circulation and auxiliary circulation.