Fiona H. Zhou

TNF‐α Mediates p38 MAP Kinase Activation and Negatively Regulates Bone Formation at the Injured Growth Plate in Rats

TNF‐α is known to inhibit osteoblast differentiation in vitro and yet it is essential for bone fracture repair. Roles of TNF‐α in the bony repair of injured growth plate were examined in young rats treated with a TNF‐α antagonist. The results show that TNF‐α mediates p38 activation, which influences the recruitment, proliferation, and osteoblast differentiation of mesenchymal cells and negatively regulates bone formation at the injured growth plate.

Expression of Bone Morphogenic Proteins and Receptors at the Injured Growth Plate Cartilage in Young Rats

The injured growth plate cartilage is often repaired by bony tissue, resulting in impaired bone growth in children. Bone morphogenic proteins (BMPs) are important for bone fracture repair, and as a step to characterize potential involvement of BMPs in bony repair of injured growth plate, expression of BMPs and receptors (BMP-R) was examined by quantitative RT-PCR and immunohistochemistry in rat injured tibial growth plate. During the inflammatory response on day 1, slightly increased expression of BMP-3, BMP-4, BMP-R1a, and BMP-R2 was observed, with immunostaining seen among inflammatory cells at the injury site. During mesenchymal infiltration and osteogenic responses on days 3-14, moderately increased expression of BMP-2, −3, −4, −7, and BMP-R1a was found, with immunostaining observed among infiltrated mesenchymal cells and differentiated osteoblasts lining bony trabeculae. During maturation phase on days 14-25, only BMP-7 was seen upregulated slightly and was localized in osteoblasts and marrow cells at the injury site. The temporospatial expression of BMPs and receptors at the injured growth plate suggests potential involvement of BMP-3 and −4 in regulating the inflammatory response or as its mediators in modulating downstream events, and BMP-2, −3, −4, and −7 in the fibrogenic and osteogenic responses, and BMP-7 in bone remodeling at the injured growth plate.

Development of Anxiety-Like Behavior via Hippocampal IGF-2 Signaling in the Offspring of Parental Morphine Exposure: Effect of Enriched Environment

Opioid addiction is a major social, economic, and medical problem worldwide. Long-term adverse consequences of chronic opiate exposure not only involve the individuals themselves but also their offspring. Adolescent maternal morphine exposure results in behavior and morphologic changes in the brain of their adult offspring. However, few studies investigate the effect of adult opiate exposure on their offspring. Furthermore, the underlying molecular signals regulating the intergenerational effects of morphine exposure are still elusive. We report here that morphine exposure of adult male and female rats resulted in anxiety-like behavior and dendritic retraction in the dentate gyrus (DG) region of the hippocampus in their adult offspring. The behavior and morphologic changes were concomitant with the downregulation of insulin-like growth factor (IGF)-2 signaling in the granular zone of DG. Overexpression of hippocampal IGF-2 by bilateral intra-DG injection of lentivirus encoding the IGF-2 gene prevented anxiety-like behaviors in the offspring. Furthermore, exposure to an enriched environment during adolescence corrected the reduction of hippocampal IGF-2 expression, normalized anxiety-like behavior and reversed dendritic retraction in the adult offspring. Thus, parental morphine exposure can lead to the downregulation of hippocampal IGF-2, which contributed to the anxiety and hippocampal dendritic retraction in their offspring. An adolescent-enriched environment experience prevented the behavior and morphologic changes in their offspring through hippocampal IGF-2 signaling. IGF-2 and an enriched environment may be a potential intervention to prevention of anxiety and brain atrophy in the offspring of parental opioid exposure.

Deletion of TRIM32 protects mice from anxiety‐ and depression‐like behaviors under mild stress

Chronic stress causes a variety of psychiatric disorders such as anxiety and depression, but its mechanism is not well understood. Tripartite motif‐containing protein 32 (TRIM32) was strongly associated with autism spectrum disorder, attention deficit hyperactivity disorder, anxiety and obsessive compulsive disorder based on a study of copy number variation, and deletion of TRIM32 increased neural proliferation and reduced apoptosis. Here, we propose that TRIM32 is involved in chronic stress‐induced affective behaviors. Using a chronic unpredictable mild stress mouse depression model, we studied expression of TRIM32 in brain tissue samples and observed behavioral changes in Trim32 knockout mice. The results showed that TRIM32 protein but not its mRNA was significantly reduced in hippocampus in a time‐dependent manner within 8 weeks of chronic stress. These stress‐induced affective behaviors and reduction of TRIM32 protein expression were significantly reversed by antidepressant fluoxetine treatment. In addition, Trim32 knockout mice showed reduced anxiety and depressive behaviors and hyperactivities compared with Trim32 wild‐type mice under normal and mild stress conditions. We conclude that TRIM32 plays important roles in regulation of hyperactivities and positively regulates the development of anxiety and depression disorders induced by chronic stress.

Mature BDNF promotes the growth of glioma cells in vitro.

High-grade glioma is incurable and is associated with a short survival time and a poor prognosis. There are two forms of brain-derived neurotrophic factor (BDNF), proBDNF and mature BDNF, which exert opposite effects. Their diverse actions are mediated through two different transmembrane receptor signalling systems: p75NTR and TrkB. The important roles of the BDNF/TrkB signalling system in tumour cell proliferation and survival have been demonstrated. However, few studies have been able to distinguish mature BDNF from proBDNF due to the limitation of specific antibodies. Using specific proBDNF antibodies, we demonstrated that the proBDNF/p75NTR pathway appears to inhibit malignant glioma cell growth and migration. In the present study using specific mature BDNF antibodies, we found that mature BDNF inhibited C6 glioma cell apoptosis and increased cell growth and migration in vitro. Our data suggest that the counterbalance between mature BDNF and proBDNF may regulate tumour growth.

Mice deficient for wild-type p53-induced phosphatase 1 display elevated anxiety- and depression-like behaviors

Mood disorders are a severe health burden but molecular mechanisms underlying mood dysfunction remain poorly understood. Here, we show that wild-type p53-induced phosphatase 1 (Wip1) negatively responds to the stress-induced negative mood-related behaviors. Specifically, we show that Wip1 protein but not its mRNA level was downregulated in the hippocampus but not in the neocortex after 4 weeks of chronic unpredictable mild stress (CUMS) in mice. Moreover, the CUMS-responsive WIP1 downregulation in the hippocampus was restored by chronic treatment of fluoxetine (i.p. 20 mg/kg) along with the CUMS procedure. In addition, Wip1 knockout mice displayed decreased exploratory behaviors as well as increased anxiety-like and depression-like behaviors in mice without impaired motor activities under the non-CUMS condition. Furthermore, the Wip1 deficiency-responsive anxiety-like but not depression-like behaviors were further elevated in mice under CUMS. Although limitations like male-alone sampling and multiply behavioral testing exist, the present study suggests a potential protective function of Wip1 in mood stabilization.

Intrathecal injection of lentivirus-mediated glial cell line-derived neurotrophic factor RNA interference relieves bone cancer-induced pain in rats

Bone cancer pain is a common symptom in cancer patients with bone metastases and the underlying mechanisms are largely unknown. The aim of this study is to explore the endogenous analgesic mechanisms to develop new therapeutic strategies for bone‐cancer induced pain (BCIP) as a result of metastases. MRMT‐1 tumor cells were injected into bilateral tibia of rats and X‐rays showed that the area suffered from bone destruction, accompanied by an increase in osteoclast numbers. In addition, rats with bone cancer showed apparent mechanical and thermal hyperalgesia at day 28 after intratibial MRMT‐1 inoculation. However, intrathecal injection of morphine or lentivirus‐mediated glial cell line‐derived neurotrophic factor RNAi (Lvs‐siGDNF) significantly attenuated mechanical and thermal hyperalgesia, as shown by increases in paw withdrawal thresholds and tail‐flick latencies, respectively. Furthermore, Lvs‐siGDNF interference not only substantially downregulated GDNF protein levels, but also reduced substance P immunoreactivity and downregulated the ratio of pERK/ERK, where its activation is crucial for pain signaling, in the spinal dorsal horn of this model of bone‐cancer induced pain. In this study, Lvs‐siGDNF gene therapy appeared to be a beneficial method for the treatment of bone cancer pain. As the effect of Lvs‐siGDNF to relieve pain was similar to morphine, but it is not a narcotic, the use of GDNF RNA interference may be considered as a new therapeutic strategy for the treatment of bone cancer pain in the future.

Injection of Anti-proBDNF in Anterior Cingulate Cortex (ACC) Reverses Chronic Stress-Induced Adverse Mood Behaviors in Mice

Foraging behavior is a species-specific behavior which is considered to involve the decision making and higher cognitive functions. We previously established a novel method to detect the foraging behavior in chronic unpredictable mild stress (CUMS)-induced depression mice, in which the food foraging activity of mice was significantly reduced. Furthermore, it is generally assumed that the bilateral anterior cingulate cortex (ACC) is related to foraging activity in rat. Brain-derived neurotrophic factor (BDNF) is widely expressed in many regions of the brain and is down-regulated in depressive patients. However, the relationship between the precursor of brain-derived neurotrophic factor (proBDNF) and depression has not been fully elucidated. The results showed that CUMS in mice induced anxiety- and depression-like behaviors and significant reduction in BDNF messenger RNA (mRNA) in the brain. In this study, we evaluated the effect of anti-BDNF and anti-proBDNF in the ACC on the CUMS-induced depression mice. In contrast to the normal IgG group (normal IgG microinjection into the ACC), bilateral ACC treatment with anti-proBDNF microinjection not only reversed depressive activity but also significantly increased the amount of foraged food and BDNF mRNA in the brain. There was no significant alteration in the group of anti-BDNF microinjection into the ACC. Our data indicate that the proBDNF signaling pathway might down-regulate the foraging activity in CUMS rodents and be involved in the depression.

Targeting prostate cancer cells with genetically engineered polypeptide-based micelles displaying gastrin-releasing peptide.

In recent years G protein-coupled receptors (GPCRs) have emerged as crucial tumorigenic factors that drive aberrant cancer growth, metastasis and angiogenesis. Consequently, a number of GPCRs are strongly expressed in cancer derived cell lines and tissue samples. Therefore a rational anti-cancer strategy is the design of nano-medicines that specifically target GPCRs to bind and internalise cytotoxic drugs into cancer cells. Herein, we report the genetic engineering of a self-assembling nanoparticle based on elastin-like polypeptide (ELP), which has been fused with gastrin releasing peptide (GRP). These nanoparticles increased intracellular calcium concentrations when added to GRP receptor positive PC-3 prostate cancer cells, demonstrating specific receptor activation. Moreover, GRP-displaying fluorescent labelled nanoparticles showed specific cell-surface interaction with PC-3 prostate cancer cells and increased endocytic uptake. These nanoparticles therefore provide a targeted molecular carrier system for evaluating the delivery of cytotoxic drugs into cancer cells.

The glucagon-like peptide-1 analogue exendin-4 reverses impaired intracellular Ca(2+) signalling in steatotic hepatocytes.

The release of Ca(2+) from the endoplasmic reticulum (ER) and subsequent replenishment of ER Ca(2+) by Ca(2+) entry through store-operated Ca(2+) channels (SOCE) play critical roles in the regulation of liver metabolism by adrenaline, glucagon and other hormones. Both ER Ca(2+) release and Ca(2+) entry are severely inhibited in steatotic hepatocytes. Exendin-4, a slowly-metabolised glucagon-like peptide-1 (GLP-1) analogue, is known to reduce liver glucose output and liver lipid, but the mechanisms involved are not well understood. The aim of this study was to determine whether exendin-4 alters intracellular Ca(2+) homeostasis in steatotic hepatocytes, and to evaluate the mechanisms involved. Exendin-4 completely reversed lipid-induced inhibition of SOCE in steatotic liver cells, but did not reverse lipid-induced inhibition of ER Ca(2+) release. The action of exendin-4 on Ca(2+) entry was rapid in onset and was mimicked by GLP-1 or dibutyryl cyclic AMP. In steatotic liver cells, exendin-4 caused a rapid decrease in lipid (half time 6.5min), inhibited the accumulation of lipid in liver cells incubated in the presence of palmitate plus the SOCE inhibitor BTP-2, and enhanced the formation of cyclic AMP. Hormone-stimulated accumulation of extracellular glucose in glycogen replete steatotic liver cells was inhibited compared to that in non-steatotic cells, and this effect of lipid was reversed by exendin-4. It is concluded that, in steatotic hepatocytes, exendin-4 reverses the lipid-induced inhibition of SOCE leading to restoration of hormone-regulated cytoplasmic Ca(2+) signalling. The mechanism may involve GLP-1 receptors, cyclic AMP, lipolysis, decreased diacylglycerol and decreased activity of protein kinase C.