Functional quantitative susceptibility mapping (fQSM) of rat brain during flashing light stimulation.

Functional magnetic resonance imaging (fMRI) based on the blood oxygenation level-dependent (BOLD) contrast has become an indispensable tool in neuroscience. However, the BOLD signal is nonlocal, lacking quantitative measurement of oxygenation fluctuation. This preclinical study aimed to introduced functional quantitative susceptibility mapping (fQSM) to complement BOLD-fMRI to quantitatively assess the local susceptibility and venous oxygen saturation (SvO2). Rats were subjected to a 5 Hz flashing light and the different inhaled oxygenation levels (30% and 100%) were used to observe the venous susceptibility to quantify SvO2. Phase information was extracted to produce QSM, and the activation responses of magnitude (conventional BOLD) and the QSM time-series were analyzed. During light stimulation, the susceptibility change of fQSM was four times larger than the BOLD signal change in both inhalation oxygenation conditions. Moreover, the responses in the fQSM map were more restricted to the visual pathway, such as the lateral geniculate nucleus and superior colliculus, compared with the relatively diffuse distributions in the BOLD map. Also, the calibrated SvO2 was approximately 84% (88%) when the task was on, 83% (87%) when the task was off during 30% (and during 100%) oxygen inhalation. This is the first fQSM study in a small animal model and increases our understanding of fQSM in the brains of small animals. This study demonstrated the feasibility, sensitivity, and specificity of fQSM using light stimulus, as fQSM provides quantitative clues as well as localized information, complementing the defects of BOLD-fMRI. In addition to neural activity, fQSM also assesses SvO2 as supplementary information while BOLD-fMRI dose not. Accordingly, the fQSM technique could be a useful quantitative tool for functional studies, such as longitudinal follow up of neurodegenerative diseases, functional recovery after brain surgery, and negative BOLD studies.

Designing a biochip following multiplex polymerase chain reaction for the detection of Salmonella serovars Typhimurium, Enteritidis, Infantis, Hadar, and Virchow in poultry products.

Salmonella-contaminated foods, especially poultry-derived foods (eggs, chicken meat), are the major source of salmonellosis. Not only in the European Union (EU), but also in the United States, Japan, and other countries, has salmonellosis been an issue of concern for food safety control agencies. In 2005, EU regulation 1003/2005 set a target for the control and reduction of five target Salmonella enterica serovars-S. Typhimurium, S. Enteritidis, S. Infantis, S. Hadar, and S. Virchow-in breeding flocks. Thus, a simple biochip for the rapid detection of any of these five Salmonella serovars in poultry products may be required. The objectives of this study were to design S. Virchow-specific primers and to develop a biochip for the simultaneous identification of all or any of these five Salmonella serovars in poultry and poultry products. Experimentally, we designed novel polymerase chain reaction (PCR) primers for the specific detection of S. Virchow, S. Infantis, and S. Hadar. The specificity of all these primers and two known primer sets for S. Typhimurium and S. Enteritidis was then confirmed under the same PCR conditions using 57 target strains and 112 nontarget Salmonella strains as well as 103 non-Salmonella strains. Following multiplex PCR, strains of any of these five Salmonella serovars could be detected by a chromogenic biochip deployed with DNA probes specific to these five Salmonella serovars. In comparison with the multiplex PCR methods, the biochip assay could improve the detection limit of each of the Salmonella serovars from N×103 cfu/mL to N×102 cfu/mL sample in either the pure culture or the chicken meat samples. With an 8-hour enrichment step, the detection limit could reach up to N×100 cfu/mL.

Detection of Salmonella in chicken meat by insulated isothermal PCR.

Consumption of Salmonella-contaminated foods, such as poultry and fresh eggs, is known to be one of the main causes of salmonellosis. Conventional PCR methods, including real-time PCR for rapid detection of Salmonella, in general require skilled technicians and costly instruments. A recently developed novel convective PCR, insulated isothermal PCR (iiPCR), is carried out in polycarbonate capillary tubes. In this study, we designed TaqMan probes and PCR primers based on the yrfH gene encoding a heat shock protein for the iiPCR detection of Salmonella in chicken meat samples. The TaqMan probe was labeled with 6-carboxyfluorescein and 6-carboxytetramethylrhodamine at the 5' and 3' ends, respectively. The PCR amplicon was 133 bp. A typical run of this iiPCR assay was completed within 1 h. Specific PCR products were obtained for 148 strains representing 49 serotypes of Salmonella tested. Under the same conditions, false-positive results were not obtained for 98 non-Salmonella strains tested, including strains of Enterobacteriaceae closely related to Salmonella. For chicken meat samples, with a 5-h enrichment step Salmonella at as low as 10° CFU/g of poultry meat could be detected. Because the amplification signals from the probes are detectable at 520 nm, identification of the PCR products by gel electrophoresis is not required. Compared with conventional PCR, the iiPCR system requires less expertise and provides an economical, reliable, and rapid tool for result interpretation. Detection results can be obtained within 8 h, including the enrichment and DNA extraction steps.

Functional and structural characteristics of tumor angiogenesis in lung cancers overexpressing different VEGF isoforms assessed by DCE- and SSCE-MRI.

The expressions of different vascular endothelial growth factor (VEGF) isoforms are associated with the degree of tumor invasiveness and the patient's prognosis in human cancers. We hypothesized that different VEGF isoforms can exert different effects on the functional and structural characteristics of tumor angiogenesis. We used dynamic contrast-enhanced MRI (DCE-MRI) and steady-state contrast-enhanced MRI (SSCE-MRI) to evaluate in vivo vascular functions (e.g., perfusion and permeability) and structural characteristics (e.g., vascular size and vessel density) of the tumor angiogenesis induced by different VEGF isoforms (VEGF121, VEGF165, and VEGF189) in a murine xenograft model of human lung cancer. Tumors overexpressing VEGF189 were larger than those overexpressing the other two VEGF isoforms. The K(trans) map obtained from DCE-MRI revealed that the perfusion and permeability functions of tumor microvessels was highest in both the rim and core regions of VEGF189-overexpressing tumors (p<0.001 for both tumor rim and core). The relative vessel density and relative vessel size indexes derived from SSCE-MRI revealed that VEGF189-overexpressing tumors had the smallest (p<0.05) and the most-dense (p<0.01) microvessels, which penetrated deeply from the tumor rim into the core, followed by the VEGF165-overepxressing tumor, whose microvessels were located mainly in the tumor rim. The lowest-density microvessels were found in the VEGF121-overexpressing tumor; these microvessels had a relatively large lumen and were found mainly in the tumor rim. We conclude that among the three VEGF isoforms evaluated, VEGF189 induces the most densely sprouting and smallest tumor microvessels with the highest in vivo perfusion and permeability functions. These characteristics of tumor microvessels may contribute to the reported adverse effects of VEGF189 overexpression on tumor progression, metastasis, and patient survival in several human cancers, including non-small cell lung cancer, and suggest that applying aggressive therapy may be necessary in human cancers in which VEGF189 is overexpressed.

Impaired water reabsorption in mice deficient in the type VI adenylyl cyclase (AC6).

Adenylyl cyclase (AC) type VI (AC6) is a calcium-inhibitable enzyme which produces cAMP upon stimulation. Herein, we characterized the specific role of AC6 in the kidneys using two AC6-knockout mouse lines. Immunohistochemical staining revealed that AC6 exists in the tubular parts of the nephron and collecting duct. Activities of AC evoked by forskolin or a selective agonist of the V2 vasopressin receptor were lower in the kidneys of AC6-null mice compared to those of wildtype mice. Results of a metabolic cage assay and dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) showed for the first time that AC6 plays a critical role in regulating water homeostasis.