Mohammed Jameel

Interdependency of β-Adrenergic Receptors and CFTR in Regulation of Alveolar Active Na+ Transport

&bgr;-Adrenergic receptors (&bgr;AR) regulate active Na+ transport in the alveolar epithelium and accelerate clearance of excess airspace fluid. Accumulating data indicates that the cystic fibrosis transmembrane conductance regulator (CFTR) is important for upregulation of the active ion transport that is needed to maintain alveolar fluid homeostasis during pulmonary edema. We hypothesized that &bgr;AR regulation of alveolar active transport may be mediated via a CFTR dependent pathway. To test this hypothesis we used a recombinant adenovirus that expresses a human CFTR cDNA (adCFTR) to increase CFTR function in the alveolar epithelium of normal rats and mice. Alveolar fluid clearance (AFC), an index of alveolar active Na+ transport, was 92% greater in CFTR overexpressing lungs than controls. Addition of the Cl− channel blockers NPPB, glibenclamide, or bumetanide and experiments using Cl− free alveolar instillate solutions indicate that the accelerated AFC in this model is due to increased Cl− channel function. Conversely, CFTR overexpression in mice with no &bgr;1- or &bgr;2-adrenergic receptors had no effect on AFC. Overexpression of a human &bgr;2AR in the alveolar epithelium significantly increased AFC in normal mice but had no effect in mice with a non-functional human CFTR gene (&Dgr;&phgr;508 mutation). These studies indicate that upregulation of alveolar CFTR function speeds clearance of excess fluid from the airspace and that CFTRs effect on active Na+ transport requires the &bgr;AR. These studies reveal a previously undetected interdependency between CFTR and &bgr;AR that is essential for upregulation of active Na+ transport and fluid clearance in the alveolus.

Association between rectal optical signatures and colonic neoplasia: potential applications for screening.

Field carcinogenesis detection represents a promising means for colorectal cancer (CRC) screening, although current techniques (e.g., flexible sigmoidoscopy) lack the requisite sensitivity. The novel optical technology low-coherence enhanced backscattering (LEBS) spectroscopy, allows identification of microscale architectural consequences of the field carcinogenesis in preclinical CRC models with unprecedented accuracy. To investigate the potential clinical translatability of this approach, we obtained biopsies from the normal-appearing rectal mucosa from patients undergoing colonoscopy (n = 219). LEBS signals were recorded through a bench-top instrument. Four parameters characterizing LEBS signal were linearly combined into a single marker. We found that LEBS signal parameters generally mirrored neoplasia progression from patients with no neoplasia, to 5 to 9 mm adenoma and to advanced adenomas. The composite LEBS marker calculated from the LEBS signal paralleled this risk status (ANOVA P < 0.001). Moreover, this was independent of CRC risk factors, benign colonic findings, or clinically unimportant lesions (diminutive adenomas, hyperplastic polyps). For advanced adenomas, the LEBS marker had a sensitivity of 100%, specificity of 80%, and area under the receiver operator characteristic curve of 0.895. Leave-one-out cross-validation and an independent data set (n = 51) supported the robustness of these findings. In conclusion, we provide the first demonstration that LEBS-detectable alterations in the endoscopically normal rectum were associated with the presence of neoplasia located elsewhere in the colon. This study provides the proof of concept that rectal LEBS analysis may potentially provide a minimally intrusive CRC screening technique. Further studies with an endoscopically compatible fiber optic probe are under way for multicenter clinical validation.

Spectroscopic Microvascular Blood Detection From the Endoscopically Normal Colonic Mucosa: Biomarker for Neoplasia Risk

BACKGROUND & AIMS We previously used a novel biomedical optics technology, 4-dimensional elastically scattered light fingerprinting, to show that in experimental colon carcinogenesis the predysplastic epithelial microvascular blood content is increased markedly. To assess the potential clinical translatability of this putative field effect marker, we characterized the early increase in blood supply (EIBS) in human beings in vivo. METHODS We developed a novel, endoscopically compatible, polarization-gated, spectroscopic probe that was capable of measuring oxygenated and deoxygenated (Dhb) hemoglobin specifically in the mucosal microcirculation through polarization gating. Microvascular blood content was measured in 222 patients from the endoscopically normal cecum, midtransverse colon, and rectum. If a polyp was present, readings were taken from the polyp tissue along with the normal mucosa 10-cm and 30-cm proximal and distal to the lesion. RESULTS Tissue phantom studies showed that the probe had outstanding accuracy for hemoglobin determination (r(2) = 0.99). Augmentation of microvasculature blood content was most pronounced within the most superficial ( approximately 100 microm) layer and dissipated in deeper layers (ie, submucosa). EIBS was detectable within 30 cm from the lesion and the magnitude mirrored adenoma proximity. This occurred for both oxygenated hemoglobin and DHb, with the effect size being slightly greater for DHb. EIBS correlated with adenoma size and was not engendered by nonneoplastic (hyperplastic) polyps. CONCLUSIONS We show, herein, that in vivo microvascular blood content can be measured and provides an accurate marker of field carcinogenesis. This technological/biological advance has numerous potential applications in colorectal cancer screening such as improved polyp detection and risk stratification.

Measuring mucosal blood supply in vivo with a polarization-gating probe

There has been significant interest in developing depth-selective optical interrogation of biological tissue in general and of superficial (e.g., mucosal) tissue in particular. We report an in vivo polarization-gating fiber-optic probe that obtains backscattering spectroscopic measurements from a range of near-surface depths (100-200 microm). The design and testing was performed with polarized light Monte Carlo simulations and in tissue model experiments. We used the probe to investigate mucosal changes in early carcinogenesis. Measurements performed in the colonic mucosa of 125 human subjects provide the first in vivo evidence that mucosal blood supply is increased early in carcinogenesis, not only in precancerous adenomatous lesions, but also in the histologically normal-appearing tissue surrounding these lesions. This effect was primarily limited to the mucosal microcirculation and was not present in the larger blood vessels located deeper in colonic tissue.

Rectal mucosal microvascular blood supply increase is associated with colonic neoplasia.

Purpose: Endoscopic examination has proven effective in both detecting and preventing colorectal cancer; however, only about a quarter of eligible patients undergo screening. Even if the compliance rate increased, limited endoscopic capacity and cost would be prohibitive. There is a need for an accurate method to target colonoscopy to those most at risk of harboring colonic neoplasia. Exploiting field carcinogenesis seems to be a promising avenue. Our group recently reported that an early increase in blood supply (EIBS) is a reliable marker of field carcinogenesis in experimental models. We now investigate whether in situ detection of EIBS in the rectum can predict neoplasia elsewhere in the colon. Experimental Design: We developed a novel polarization-gated spectroscopy fiber-optic probe that allows depth-selective interrogation of microvascular blood content. Using the probe, we examined the blood content in vivo from the rectal mucosa of 216 patients undergoing screening colonoscopy. Results: Microvascular blood content was increased by ∼50% in the endoscopically normal rectal mucosa of patients harboring advanced adenomas when compared with neoplasia-free patients irrespective of lesion location. Demographic factors and nonneoplastic lesions did not confound this observation. Logistic regression using mucosal oxyhemoglobin concentration and patient age resulted in a sensitivity of 83%, a specificity of 82%, and an area under the receiver operating characteristic curve of 0.88 for the detection of advanced adenomas. Conclusions: Increased microvascular blood supply in the normal rectal mucosa is associated with the presence of clinically significant neoplasia elsewhere in the colon, supporting the development of rectal EIBS as a colon cancer risk-stratification tool.

In vivo timing of onset of transgene expression following adenoviral-mediated gene transfer.

Recombinant adenoviruses are efficient gene transfer vehicles that could be used for treatment of acute diseases. However, the time required for adenoviruses to produce physiologically relevant levels of transgene in vivo is unknown. To address this question rat lungs were infected with an E1a(-)/E3a(-) adenovirus that contains an hCMV-driven human beta(2)-adrenergic receptor (beta(2)AR) cDNA. Human beta(2)AR message and protein expression were noted 2-4 h postinfection without evidence of pseudotransduction. beta(2)AR function (cAMP production) was increased at 6 h postinfection. To determine when beta(2)AR gene transfer affects downstream catecholamine-sensitive pathways, we measured lung Na,K-ATPase expression and alveolar fluid clearance (AFC). beta(2)AR gene transfer increased Na,K-ATPase number by 80% at 6 h, and AFC by 20% at 8 h postinfection. These data indicate that recombinant adenoviruses can produce physiologically significant levels of transgene within hours of infection and that they may be suitable for gene therapies for acute, rapidly progressive diseases.

Spectral Slope from the Endoscopically-Normal Mucosa Predicts Concurrent Colonic Neoplasia: A Pilot Ex-Vivo Clinical Study

PurposeWe previously reported that analysis of histologically normal intestinal epithelium for spectral slope, a marker for aberrations in nanoscale tissue architecture, had outstanding accuracy in identifying field carcinogenesis in preclinical colorectal cancer models. In this study, we assessed the translatability of spectral slope analysis to human colorectal cancer screening.MethodsSubjects (n = 127) undergoing colonoscopy had spectral slope determined from two endoscopically normal midtransverse colonic biopsies using four-dimensional elastic light-scattering fingerprinting and correlated with clinical findings.ResultsFour-dimensional elastic light-scattering fingerprinting analysis showed the submicron particles size progressively shifted toward larger sizes in subjects harboring neoplasia. There was a corresponding decrease in spectral slope values from the endoscopically normal mucosa in subjects harboring adenomas (n = 41) and advanced adenomas (n = 10), compared to neoplasia-free subjects (P ≤ 0.00001). These factors did not appear to be confounded by either age or adenoma location. For detecting advanced adenomas, spectral slope had a negative and positive predictive value of 95 percent and 50 percent respectively.ConclusionsWe demonstrate, for the first time, that spectral slope in “normal” mucosa can accurately risk-stratify patients for colonic neoplasia. This proof of concept study serves to underscore the promise of four-dimensional elastic light-scattering fingerprinting analysis for colorectal cancer screening.