William Mustain

Inhibition of Fatty Acid Synthase Attenuates CD44-Associated Signaling and Reduces Metastasis in Colorectal Cancer

Fatty acid synthase (FASN) and ATP-citrate lyase, key enzymes of de novo lipogenesis, are significantly upregulated and activated in many cancers and portend poor prognosis. Even though the role of lipogenesis in providing proliferative and survival advantages to cancer cells has been described, the impact of aberrant activation of lipogenic enzymes on cancer progression remains unknown. In this study, we found that elevated expression of FASN is associated with advanced stages of colorectal cancer (CRC) and liver metastasis, suggesting that it may play a role in progression of CRC to metastatic disease. Targeted inhibition of lipogenic enzymes abolished expression of CD44, a transmembrane protein associated with metastases in several cancers including CRC. In addition, inhibition of lipogenic enzymes and reduced expression of CD44 attenuated the activation of MET, Akt, FAK, and paxillin, which are known to regulate adhesion, migration, and invasion. These changes were consistent with an observed decrease in migration and adhesion of CRC cells in functional assays and with reorganization of actin cytoskeleton upon FASN inhibition. Despite the modest effect of FASN inhibition on tumor growth in xenografts, attenuation of lipogenesis completely abolished establishment of hepatic metastasis and formation of secondary metastasis. Together, our findings suggest that targeting de novo lipogenesis may be a potential treatment strategy for advanced CRC.

Vascular endothelium growth factor, surgical delay, and skin flap survival.

Objective:Cytokines may be a mechanism by which surgical delay can increase flap survival. We previously found that preoperative vascular endothelium growth factor (VEGF) administration in the rat transverse rectus abdominis myocutaneous (TRAM) flap could improve skin paddle survival. In this study, we used partial elevation of the rat TRAM flap as a surgical delay to assess endogenous cytokine expression and tissue survival comparable to undelayed TRAM flaps. Methods:In Part I, TRAM flaps underwent surgical delay procedures; 7 days later, the flaps were completely elevated and reinset. At the same time, other flaps were raised and reinset without delay. Skin paddle survival in both groups was evaluated at 7 days. In Part II, skin biopsies from TRAM zones I to IV were taken at the time of delay and at intervals of 12, 24, 48, and 72 hours. Specimens were assessed for selected cytokine gene expression by reverse transcription-polymerase chain reaction analysis (TR-PCR). Results:Surgical delay significantly (P < 0.001) increased skin paddle survival in the delayed TRAM flaps (16.14 ± 1.53 cm, 81.9%) compared with undelayed flaps (7.68 ± 3.16 cm, 40.9%). TGF-&bgr; and PDGF expressions were not changed by surgical delay, but basic fibroblast growth factor (bFGF) and VEGF expressions increased significantly (P < 0.05 and P < 0.01) after delay. Conclusions:In the rat TRAM model, surgical delay resulted in increased VEGF expression and increased skin paddle survival. These results correlate with previous studies showing the preoperative injection of VEGF increases skin paddle survival. VEGF may be an important element in the delay phenomenon and may be an agent for pharmacological delay.

The role of neurotensin in physiologic and pathologic processes.

Purpose of reviewNeurotensin is a 13-amino acid peptide found in the central nervous system central nervous system and the gastrointestinal tract. Since its initial discovery in 1973, neurotensin has been shown to play a role in a wide range of physiologic and pathologic processes throughout the body. Ongoing research efforts continue to clarify the role of neurotensin in various central nervous system and gastrointestinal processes, as well as how disruption of these normal mechanisms may lead to diseases ranging from schizophrenia to colorectal cancer. The goal of this review is to provide an overview of the most recent advances in the field of neurotensin research, in the context of what has been previously published. Recent findingsBecause of the seemingly unrelated functions of neurotensin in the central nervous system and the periphery, the scope of the articles reviewed is rather broad. Contributions continue to be made to our understanding of the downstream effects of neurotensin signaling and the complex feedback loops between neurotensin and other signaling molecules. By selective targeting or blockade of specific neurotensin receptors, investigators have identified potential drugs for use in the treatment of schizophrenia, alcoholism, chronic pain, or cancer. Neurotensin-based pharmacologic agents are being used successfully in animal models for a number of these conditions. SummaryThe review highlights the wide array of biological processes in which neurotensin has a role, and summarizes the most recent advances in various fields of neurotensin research. The knowledge gained through this research has led to the development of first-in-class drugs for the treatment of various medical conditions, and it is clear that in the coming years some of these agents will be ready to move from the bench to the bedside in clinical trials.

Delivery of RNA Nanoparticles into Colorectal Cancer Metastases Following Systemic Administration

The majority of deaths from all cancers, including colorectal cancer (CRC), is a result of tumor metastasis to distant organs. To date, an effective and safe system capable of exclusively targeting metastatic cancers that have spread to distant organs or lymph nodes does not exist. Here, we constructed multifunctional RNA nanoparticles, derived from the three-way junction (3WJ) of bacteriophage phi29 motor pRNA, to target metastatic cancer cells in a clinically relevant mouse model of CRC metastasis. The RNA nanoparticles demonstrated metastatic tumor homing without accumulation in normal organ tissues surrounding metastatic tumors. The RNA nanoparticles simultaneously targeted CRC cancer cells in major sites of metastasis, such as liver, lymph nodes, and lung. Our results demonstrate the therapeutic potential of these RNA nanoparticles as a delivery system for the treatment of CRC metastasis.

Click-evoked responses in vestibular afferents in rats

Sound activates not only the cochlea but also the vestibular end organs. Research on this phenomenon led to the discovery of the sound-evoked vestibular myogenic potentials recorded from the sternocleidomastoid muscles (cervical VEMP, or cVEMP). Since the cVEMP offers simplicity and the ability to stimulate each labyrinth separately, its values as a test of human vestibular function are widely recognized. Currently, the cVEMP is interpreted as a test of saccule function based on the assumption that clicks primarily activate the saccule. However, sound activation of vestibular end organs other than the saccule has been reported. To provide the neural basis for interpreting clinical VEMP testing, we employed the broadband clicks used in clinical VEMP testing to examine the sound-evoked responses in a large sample of vestibular afferents in Sprague-Dawley rats. Recordings were made from 924 vestibular afferents from 106 rats: 255 from the anterior canal (AC), 202 from the horizontal canal (HC), 177 from the posterior canal (PC), 207 from the superior vestibular nerve otolith (SO), and 83 from the inferior nerve otolith (IO). Sound sensitivity of each afferent was quantified by computing the cumulative probability of evoking a spike (CPE). We found that clicks activated irregular afferents (normalized coefficient of variation of interspike intervals >0.2) from both the otoliths (81%) and the canals (43%). The order of end organ sound sensitivity was SO = IO > AC > HC > PC. Since the sternocleidomastoid motoneurons receive inputs from both the otoliths and the canals, these results provide evidence of a possible contribution from both of them to the click-evoked cVEMP.

Alexander's Law Revisited

Alexanders Law states that the slow-phase velocity of the nystagmus caused by unilateral vestibular lesion increases with gaze in the beat direction. Two studies have shown that this gaze effect is generalized to the nystagmus caused by unilateral cold water irrigation. This indicates that the gaze effect is not the result of central changes associated with a peripheral lesion but rather because of unilateral vestibular peripheral inhibition. In this study, we show that there is a similar gaze effect on the nystagmus produced by unilateral warm water ear irrigation. Furthermore, we examined the two hypotheses of Alexanders Law proposed in the two studies. One hypothesis is based on the gaze-dependent modulation of the vestibulo-ocular reflex (VOR) response to unbalanced canal input. The other hypothesis, however, is based on the leaky neural integrator caused by unilateral vestibular peripheral inhibition. These two hypotheses predict the same gaze effect on the nystagmus produced by cold water irrigation, but opposite gaze effects on the nystagmus produced by warm water irrigation. Our results support the first hypothesis and suggest that the second hypothesis needs to be modified.

Improvement of venous flap survival by application of vascular endothelial growth factor in a rat model.

Expression of vascular endothelial growth factor (VEGF) in the venous flap and the effect of exogenous VEGF on survival of the venous flap were studied in rats. A 4- × 4-cm groin type 2 venous skin flap was used in the study. In part 1, biopsies were taken from the flap at 0, 6, 12, 24, and 48 hours after the flaps were raised. VEGF gene expression was measured. In part 2, exogenous VEGF (1 μg/mL) was injected subdermally into the flaps in 10 rats before the flaps were replaced. Flaps that received a saline injection were used as the control. Skin paddle survival was measured on postoperative day 7. The results showed that VEGF expression was significantly increased at 24 and 48 hours after venous flap elevation (P < 0.05). Injection of exogenous VEGF to the flap significantly improved survival of the flap (73% of the flap) when compared with the control, which had a 39% mean percent survival (P < 0.05). We conclude that VEGF expression was increased in the venous flap. Administration of exogenous VEGF significantly improved survival of the venous flap.

Frequency tuning of bone-conducted tone burst-evoked myogenic potentials recorded from extraocular muscles (BOVEMP) in normal human subjects.

In this study, we characterized the frequency tuning of bone‐conducted sound‐evoked myogenic potentials recorded from extraocular muscles (BOVEMP) in normal human subjects.

Prefabrication of vascularized bone flap by demineralized bone matrix.

Demineralized bone matrix (DBM) has been reported to have osteoconductive and osteoinductive properties and has been clinically used as a bone graft alternative. In the present study we attempted to generate a vascularized bone flap by subcutaneous implantation of DBM with a vascular loop to provide blood supply in a rat model. Thirty male Sprague-Dawley rats were divided into two groups according to the presence or absence of blood supply. In the experimental group, the bone flap was created by application of 0.4 mL of DBM onto two pieces of gelatin sponge sheets between which a vascular loop was sandwiched. A prefabricated flap without a vascular loop served as the control. The flaps were biopsied at three different time intervals postoperatively (2, 4, and 6 weeks). The results showed that DBM induced subcutaneous bone formation in both of the groups. However, in the nonvascularized group, the amount of bony tissue had decreased at four postoperative weeks and continued to do so afterwards. In contrast, bone formation was active at four weeks in the vascularized group. Our study indicated that implantation of DBM can prefabricate a bone flap. Blood supply to the flap is considered a key factor of the success of this prefabrication.

Fluorescein as a topical fluorescent contrast agent for quantitative microendoscopic inspection of colorectal epithelium

Fiber bundle microendoscopic imaging of colorectal tissue has shown promising results, for both qualitative and quantitative analysis. A quantitative image quality control and image feature extraction algorithm was previously designed for quantitative image feature analysis of proflavine-stained ex vivo colorectal tissue. We investigated fluorescein as an alternative topical stain. Images of ex vivo porcine, caprine, and human colorectal tissue were used to compare microendoscopic images of tissue topically stained with fluorescein and proflavine solutions. Fluorescein was shown to be comparable for automated crypt detection, with an average crypt detection sensitivity exceeding 90% using a combination of three contrast limit pairs.