Structured-light surface scanning system to evaluate breast morphology in standing and supine positions.

Breast shapes are affected by gravitational loads and deformities. Measurements obtained in the standing position may not correlate well with measurements in the supine position, which is more representative of patient position during breast surgery. A dual color 3D surface imaging system capable of scanning patients in both supine and standing positions was developed to evaluate the effect of changes in body posture on breast morphology. The system was evaluated with  breast phantoms to assess accuracy, then tested on ten subjects in three body postures to assess its effectiveness as a clinical tool. The accuracy of the system was within 0.4 mm on average across the model. For the human study, there was no effect of body posture on breast volumes (p value > 0.05), but we observed an effect of completeness of breast scans on body posture (p value < 0.05). Post-hoc tests showed that the supine position and the standing position with hands at the waist differed significantly (p value < 0.05). This study shows that the system can quantitatively evaluate the effect of subject postures, and thereby has the potential to be used to investigate peri-operative changes in breast morphology.

Lipid-weighted intraoperative photoacoustic tomography of breast tumors: Volumetric comparison to preoperative MRI.

With a lifetime risk of 1 in 8, breast cancer continues to be a major concern for women and their physicians. The optimal treatment of the disease depends on the stage of the cancer at diagnosis, which is typically assessed using medical imaging. However, currently employed imaging systems for breast tumor measurement rarely agree perfectly. Our group developed an Intraoperative Photoacoustic Screening (iPAS) soft tissue scanner featuring high bulk tissue sensitivity, a clinically compatible scan-time of 6 min, imaging depths greater than 2 cm and the capability to visualize whole breast tumors based on their lipid, rather than hemoglobin, profile. Here, we report on the first clinical experience with breast cancer patients by comparing tumor-measurement using iPAS, preoperative dynamic contrast enhanced magnetic resonance imaging (DCE-MRI) and gold-standard pathology. Tumor size was measured volumetrically for iPAS and DCE-MRI, and separately using maximum diameters for pathology, DCE-MRI and iPAS. Comparisons were performed using Pearson's correlation coefficients, and the non-parametric Wilcoxon signed-rank test. Twelve consecutive patients were included in the study, contingent on pathologically documented invasive carcinoma. iPAS volumetric tumor size was positively correlated to DCE-MRI (Pearson's r = 0.78, p = 0.003) and not significantly different (Wilcoxon, p = 0.97). In comparison to pathology, tumor diameters given by iPAS were positively correlated (Pearson's r = 0.87, p = 0.0002) and significantly different (Wilcoxon, p = 0.0015). The results indicated that volumetric-measurement of invasive breast tumors with iPAS is similar to that of DCE-MRI. On the other hand, tumor diameter measurements were less reliable. Beyond enhancing surgical specimen examination, an extension of this technology to diagnostic imaging promises a new perspective on tumor assessment, potentially improving our current understanding and treatment of breast cancer.

Intraoperative photoacoustic screening of breast cancer: a new perspective on malignancy visualization and surgical guidance.

High re-excision rates in breast-conserving surgery call for a new intraoperative approach to the lumpectomy margin evaluation problem. The unique intraoperative imaging system, presented here, demonstrated the capability of photoacoustic tomography (PAT) to deliver optical sensitivity and specificity, along with over 2-cm imaging depth, in a clinical setting. The system enabled the evaluation of tumor extent, shape, morphology, and position within lumpectomy specimens measuring up to 11 cm in diameter. The investigation included all major breast cancer-related lesions, such as invasive ductal carcinoma (IDC), multifocal IDC, ductal carcinoma in situ and combinations of these variants. Coregistration with established ultrasound (US) technology, as well as comparison to specimen radiography, validated the performance of PAT, which appeared to facilitate better tumor visualization. Contrary to expected PA contrast mechanisms, PAT images of hemoglobin distribution correlated poorly with US-determined tumor location, while hypointense regions in lipid-weighted PAT images were in better agreement with US.

Direct comparison of the abilities of bone marrow mesenchymal versus hematopoietic stem cells to reverse hyperglycemia in diabetic NOD.SCID mice.

Both bone marrow-derived hematopoietic stem cells (HSC) and mesenchymal stem cells (MSC) improve glycemic control in diabetic mice, but their kinetics and associated changes in pancreatic morphology have not been directly compared. Our goal was to examine the time course of improvements in glucose tolerance and associated changes in ß-cell mass and proliferation following transplantation of equivalent numbers of HSC or MSC from the same bone marrow into diabetic non-obese diabetic severe combined immune deficiency (NOD.SCID) mice. We used transgenic mice with a targeted expression of yellow fluorescent protein (YFP) driven by the Vav1 gene promoter to genetically tag HSC and progeny. HSC were separated from bone marrow by fluorescence-activated cell sorting and MSC following cell culture. Equivalent numbers of isolated HSC or MSC were transplanted directly into the pancreas of NOD.SCID mice previously made diabetic with streptozotocin. Glucose tolerance, serum insulin, ß-cell mass and ß-cell proliferation were examined up to 28 days following transplant. Transplantation with MSC improved glucose tolerance within 7 days and serum insulin levels increased, but with no increase in ß-cell mass. Mice transplanted with HSC showed improved glucose tolerance only after 3 weeks associated with increased ß-cell proliferation and mass. We conclude that single injections of either MSC or HSC transiently improved glycemic control in diabetic NOD.SCID mice, but with different time courses. However, only HSC infiltrated the islets and were associated with an expanded ß-cell mass. This suggests that MSC and HSC have differing mechanisms of action.

Mesoscopic reflectance angular domain spectroscopic imaging.

The advancement of angular domain imaging in mesoscopic reflectance multispectral imaging is reported. The key component is an angular filter array that performs the angular filtration of the back-scattered photons and generates image contrast due to the variances in tissue optical properties. The proposed modality enables multispectral imaging of subsurface features for samples too thick for transillumination angular domain spectroscopic imaging (ADSI) approaches. The validation was carried out with tissue-mimicking phantoms with multiple absorptive features embedded below the surface. Multispectral images in the range of 666 to 888 nm clearly revealed the location of the features with the background scattering levels up to 20 cm⁻¹. The shape of the features was recoverable at depths of up to three to four times the transport mean free path. The spatial resolution was <1 mm and the field-of-view was larger than 2.5 cm x 30. cm. Furthermore, the attenuation spectra of measured absorptive features were successfully extracted. Target detectability and imaging quality with different background scattering levels, target depths, and illumination focal depths were discussed, as well as the capability of ADSI in reflectance optical mesoscopic imaging and its potential applications.

Postnatal development of the endocrine pancreas in mice lacking functional GABAB receptors.

Adult mice lacking functional GABAB receptors (GABAB1KO) have glucose metabolism alterations. Since GABAB receptors (GABABRs) are expressed in progenitor cells, we evaluated islet development in GABAB1KO mice. Postnatal day 4 (PND4) and adult, male and female, GABAB1KO, and wild-type littermates (WT) were weighed and euthanized, and serum insulin and glucagon was measured. Pancreatic glucagon and insulin content were assessed, and pancreas insulin, glucagon, PCNA, and GAD65/67 were determined by immunohistochemistry. RNA from PND4 pancreata and adult isolated islets was obtained, and Ins1, Ins2, Gcg, Sst, Ppy, Nes, Pdx1, and Gad1 transcription levels were determined by quantitative PCR. The main results were as follows: 1) insulin content was increased in PND4 GABAB1KO females and in both sexes in adult GABAB1KOs; 2) GABAB1KO females had more clusters (<500 µm(2)) and less islets than WT females; 3) cluster proliferation was decreased at PND4 and increased in adult GABAB1KO mice; 4) increased ß-area at the expense of the α-cell area was present in GABAB1KO islets; 5) Ins2, Sst, and Ppy transcription were decreased in PND4 GABAB1KO pancreata, adult GABAB1KO female islets showed increased Ins1, Ins2, and Sst expression, Pdx1 was increased in male and female GABAB1KO islets; and 6) GAD65/67 was increased in adult GABAB1KO pancreata. We demonstrate that several islet parameters are altered in GABAB1KO mice, further pinpointing the importance of GABABRs in islet physiology. Some changes persist from neonatal ages to adulthood (e.g., insulin content in GABAB1KO females), whereas other features are differentially regulated according to age (e.g., Ins2 was reduced in PND4, whereas it was upregulated in adult GABAB1KO females).

Susceptibility to fatty acid-induced ß-cell dysfunction is enhanced in prediabetic diabetes-prone biobreeding rats: a potential link between ß-cell lipotoxicity and islet inflammation.

ß-Cell lipotoxicity is thought to play an important role in the development of type 2 diabetes. However, no study has examined its role in type 1 diabetes, which could be clinically relevant for slow-onset type 1 diabetes. Reports of enhanced cytokine toxicity in fat-laden islets are consistent with the hypothesis that lipid and cytokine toxicity may be synergistic. Thus, ß-cell lipotoxicity could be enhanced in models of autoimmune diabetes. To determine this, we examined the effects of prolonged free fatty acids elevation on ß-cell secretory function in the prediabetic diabetes-prone BioBreeding (dp-BB) rat, its diabetes-resistant BioBreeding (dr-BB) control, and normal Wistar-Furth (WF) rats. Rats received a 48-h iv infusion of saline or Intralipid plus heparin (IH) (to elevate free fatty acid levels ~2-fold) followed by hyperglycemic clamp or islet secretion studies ex vivo. IH significantly decreased ß-cell function, assessed both by the disposition index (insulin secretion corrected for IH-induced insulin resistance) and in isolated islets, in dp-BB, but not in dr-BB or WF, rats, and the effect of IH was inhibited by the antioxidant N-acetylcysteine. Furthermore, IH significantly increased islet cytokine mRNA and plasma cytokine levels (monocyte chemoattractant protein-1 and IL-10) in dp-BB, but not in dr-BB or WF, rats. All dp-BB rats had mononuclear infiltration of islets, which was absent in dr-BB and WF rats. In conclusion, the presence of insulitis was permissive for IH-induced ß-cell dysfunction in the BB rat, which suggests a link between ß-cell lipotoxicity and islet inflammation.

Transillumination hyperspectral imaging for histopathological examination of excised tissue.

Angular domain spectroscopic imaging (ADSI) is a novel technique for the detection and characterization of optical contrast in turbid media based on spectral characteristics. The imaging system employs a silicon micromachined angular filter array to reject scattered light traversing a specimen and an imaging spectrometer to capture and discriminate the largely remaining quasiballistic light based on spatial position and wavelength. The imaging modality results in hyperspectral shadowgrams containing two-dimensional (2D) spatial maps of spectral information. An ADSI system was constructed and its performance was evaluated in the near-infrared region on tissue-mimicking phantoms. Image-based spectral correlation analysis using transmission spectra and first order derivatives revealed that embedded optical targets could be resolved. The hyperspectral images obtained with ADSI were observed to depend on target concentration, target depth, and scattering level of the background medium. A similar analysis on a muscle and tumor sample dissected from a mouse resulted in spatially dependent optical transmission spectra that were distinct, which suggested that ADSI may find utility in classifying tissues in biomedical applications.

Disruption of the dopamine d2 receptor impairs insulin secretion and causes glucose intolerance.

The relationship between antidopaminergic drugs and glucose has not been extensively studied, even though chronic neuroleptic treatment causes hyperinsulinemia in normal subjects or is associated with diabetes in psychiatric patients. We sought to evaluate dopamine D2 receptor (D2R) participation in pancreatic function. Glucose homeostasis was studied in D2R knockout mice (Drd2(-/-)) mice and in isolated islets from wild-type and Drd2(-/-) mice, using different pharmacological tools. Pancreas immunohistochemistry was performed. Drd2(-/-) male mice exhibited an impairment of insulin response to glucose and high fasting glucose levels and were glucose intolerant. Glucose intolerance resulted from a blunted insulin secretory response, rather than insulin resistance, as shown by glucose-stimulated insulin secretion tests (GSIS) in vivo and in vitro and by a conserved insulin tolerance test in vivo. On the other hand, short-term treatment with cabergoline, a dopamine agonist, resulted in glucose intolerance and decreased insulin response to glucose in wild-type but not in Drd2(-/-) mice; this effect was partially prevented by haloperidol, a D2R antagonist. In vitro results indicated that GSIS was impaired in islets from Drd2(-/-) mice and that only in wild-type islets did dopamine inhibit GSIS, an effect that was blocked by a D2R but not a D1R antagonist. Finally, immunohistochemistry showed a diminished pancreatic beta-cell mass in Drd2(-/-) mice and decreased beta-cell replication in 2-month-old Drd2(-/-) mice. Pancreatic D2Rs inhibit glucose-stimulated insulin release. Lack of dopaminergic inhibition throughout development may exert a gradual deteriorating effect on insulin homeostasis, so that eventually glucose intolerance develops.

Exposure of the pregnant rat to low protein diet causes impaired glucose homeostasis in the young adult offspring by different mechanisms in males and females.

The understanding of the mechanisms by which gender dimorphisms are involved in the modulation of insulin sensitivity and glucose tolerance can be crucial to unravel the development of type 2 diabetes. Rats treated with a low protein diet (LP, 8% protein content) during pregnancy and lactation have a reduced beta-cell mass at birth and a reduced insulin secretion at weaning. In this study we examined the effect of LP diet on glucose homeostasis from birth to adulthood when offspring previously exposed to LP were subsequently switched to control diet (C, 20% protein content) at weaning. The LP group had a reduced body weight after weaning compared to the C-fed rats, although their food intake was not significantly different. Furthermore, LP males had a significant increase in visceral adiposity relative to their body weight (P < 0.05). Intraperitoneal glucose tolerance test (IGTT) showed that glucose clearance was unchanged until 130 days of age when LP-fed females showed elevated blood glucose compared to C, despite similar plasma insulin levels. Females also demonstrated a significant reduction in mean pancreatic islet number, individual islet size and beta cell mass. However, no differences in IGTT or islet morphometry were observed in LP males, although basal insulin levels were twofold higher. Akt phosphorylation in response to insulin was reduced in adipose and skeletal muscle of adult rats following exposure to LP diet in early life when compared to control-fed animals, but this was only apparent in males. Plasma testosterone levels were also reduced in males at 130 days age. These data suggest that the development of impaired glucose homeostasis in offspring of LP-fed rats is likely to occur by different mechanisms in males and females.

Depth of photothermal conversion of gold nanorods embedded in a tissue-like phantom.

Gold nanorod (AuNR)-assisted photothermal therapy has emerged as a viable method for selective killing of cancer cells and shows promise for tumor destruction in vivo. This study examined the distribution of AuNR conversion expected to occur during photothermal therapy in vivo. Tissue-like phantoms were prepared with polyethylene glycol AuNRs distributed homogeneously at a concentration representative of a systemic injection. Phantoms were illuminated with a nanosecond pulsed laser (800 nm) at a variety of combinations of pulse energy (12-120 mJ) and pulse count (1-1000). Operating at the American National Standards Institute safety limit for human skin exposure (30 mJ cm(-2)), a diameter of 13 mm and a depth of 7.6 mm of AuNR conversion were observed in the gel phantoms after 1000 laser pulses (100 s exposure). Significant AuNR conversion was measured to a depth of 6 mm after only 100 pulses. Comparison of the measured AuNR conversion distribution with Monte Carlo simulation suggested that the fluence threshold for AuNR conversion estimated from phantom measurements was in the range of 20-43 mJ cm(-2). The results suggest that AuNR-assisted photothermal therapy will be effective for tumors within 10 mm of the illuminated tissue surface.

A low protein diet in early life delays the onset of diabetes in the non-obese diabetic mouse.

Dietary insult in early life can affect the development and future function of the endocrine pancreas. We maintained pregnant non-obese diabetic (NOD) mice on a low protein (LP, 8% protein versus control, 20%) diet from conception until the weaning of pups at day 21. Serum insulin and pancreatic insulin content were reduced in LP-fed NOD offspring at 8 weeks, as were serum interferon gamma and pancreatic tumor necrosis factor alpha, while the number of pancreatic islets demonstrating peri-insulitis, and the degree of invasiveness were reduced. To determine if LP caused early morphometric changes in the pancreas, we measured mean islet area at days 3 and 21. Mean islet size did not differ with diet, but by 8 weeks of age LP-fed NOD females exhibited a significantly reduced islet number and mean islet area, and a lower fractional area of pancreas occupied by both alpha- and beta-cells than control-fed mice. The onset of diabetes was delayed in NOD mice of both genders fed LP diet. The mechanism is likely to involve both altered beta-cell morphology and function and changes in cytotoxic cytokines.

Altered pancreatic morphology in the offspring of pregnant rats given reduced dietary protein is time and gender specific.

Restriction of dietary protein during gestation and lactation in the rat results in a reduction in beta cell mass, insulin content and release in the offspring, and glucose intolerance when the offspring reach adulthood. The present study was designed to identify if a particular developmental window existed during prenatal development when endocrine pancreatic development was most susceptible to nutritional insult. Pregnant rats received a low-protein (8%, LP), but isocalorific diet from conception to parturition, during the first 2 weeks of gestation (LP (1-2)), the second week only (LP (2)), or the third week (LP (3)). At other times, they received a 20% protein (C) diet, while control animals received this diet continuously. When the offspring were examined at 130 days age, animals that had received LP diet had a significantly impaired glucose tolerance compared with control-fed animals. Pancreatic morphology was examined in the offspring on postnatal days 1 and 21. The LP diet resulted in a significant decrease in the numbers of large (more than 10 000 microm(2)) and medium (between 5000 and 10 000 microm(2)) sized islets present at postnatal day 1 for all LP treatments. Consequently, mean islet area and the mean number of beta cells were reduced. The impact of LP diet was most pronounced in LP (2) for females and in LP (3) for males, and this was greater than for continuous LP exposure. Insulin and Glut-2 mRNA expression were impacted negatively by LP in early and late gestation, but increased following administration in mid-gestation. Total pancreatic insulin content was not altered by LP treatment. Pdx-1, a transcription factor associated with both beta cell development and insulin gene transcription, was decreased in female offspring following LP (1-2) and LP (3), but not in males. Pancreatic expression of nestin mRNA, and the abundance of nestin-immunoreactive cells within islets, was decreased by all LP treatments. By postnatal day 21, the mean islet area and number of beta cells had largely recovered. However, insulin and Glut-2 mRNAs were elevated in offspring exposed to LP diet, particularly in females. The studies show that LP dietary insult in early, middle, or late gestation, all result in a relative deficiency of beta cells following birth, due to a failure to develop larger islets, but that females were particularly susceptible in mid-gestation and males in late gestation.

Adipose tissue gene expression profiling reveals distinct molecular pathways that define visceral adiposity in offspring of maternal protein-restricted rats.

There is increasing evidence that poor early growth confers an increased risk of type 2 diabetes, hypertension, and other features of the metabolic syndrome in later life. We hypothesized that this may result from poor nutrition during early life exerting permanent effects on the structure and function of key metabolic organ systems. To study the long-term impact of early-life undernutrition on susceptibility to visceral adiposity, we used a rat model of maternal protein restriction (MPR) in which dams were fed a low-protein diet (containing 8% instead of 20% protein in control diet) throughout pregnancy and lactation. MPR offspring were born smaller than controls (offspring of dams on control diet) and in adulthood developed visceral adiposity. We compared the pattern of gene expression in visceral adipose tissue (VAT) between MPR offspring and controls with Affymetrix rat expression arrays. Of the total number of genes and expressed sequence tags analyzed (15,923 probe sets), 9,790 (61.5%) were expressed in VAT. We identified 650 transcripts as differentially expressed > or =1.5-fold in the VAT of MPR offspring. Gene ontology analysis revealed a global upregulation of genes involved in carbohydrate, lipid, and protein metabolism. A number of genes involved in adipocyte differentiation, angiogenesis, and extracellular matrix remodeling were also upregulated. However, in marked contrast to other rodent models of obesity, the expression of a large number of genes associated with inflammation was reduced in this rat model. Thus visceral adiposity in this early-life programmed rat model is marked by dynamic changes in the transcriptional profile of VAT. Our data provide new insights into the molecular mechanisms that underlie the early-life programming of visceral adiposity.

Gonadotropin-releasing hormone signaling pathways in an experimental ovarian tumor.

Previous results showed that GnRH signaling is altered in cells from rat luteinized ovarian tumors (tumor group) because it did not activate the phospholipase C pathway, in contrast to control ovarian cells from superovulated prepubertal rats (SPO). In the present work, alternate GnRH-induced second messengers such as phospholipase A(2) and phospholipase D activation, cAMP production, ERK1/2 phosphorylation, and the presence of G proteins were evaluated to determine GnRH mechanism of action in tumor cells. G proteins examined were present in both cell types. Buserelin, a GnRH agonist, (1, 10, and 100 ng/ml) increased phosphatidylethanol in SPO, indicating phospholipase D activation. Only 100 ng/ml buserelin induced a significant response in the tumor group. Buserelin (100 ng/ml) increased (3)H-arachidonic acid in culture media in SPO, indicating phospholipase A(2) activation; no effect was observed in the tumor group. Buserelin (100 and 1000 ng/ml) induced pertussis toxin-insensitive cAMP increases in both cell types, with similar potencies. In the tumor group, buserelin (100 ng/ml) inhibited human chorionic gonadotropin-induced cAMP and progesterone; this effect was protein kinase C (PKC) dependent (inhibited by GF109203X, a PKC inhibitor). Buserelin (100 and 1000 ng/ml) induced ERK1/2 phosphorylation in both cell kinds. Buserelin-induced ERK1/2 activation was G(i/0) independent and PKC dependent. Only in the tumor group, buserelin-induced ERK1/2 activation was cAMP dependent (abolished by SQ 22536, the adenylyl cyclase inhibitor). Furthermore, dibutyryl cAMP-induced ERK1/2 activation in the tumor group was PKC dependent (inhibited by GF109203X). In conclusion, activation of phospholipases in tumor cells does not seem to mediate GnRH effects. GnRH signaling seems to involve adenylyl cyclase activation, PKC stimulation, and ERK1/2 phosphorylation.