Contrast enhanced ultrasound imaging can predict vascular-targeted photodynamic therapy induced tumor necrosis in small animals.

AIMS: To evaluate the accuracy of contrast-enhanced ultrasound (CEUS) for monitoring tumor necrosis following WST-11 vascular targeted photodynamic therapy (VTP) using imaging-pathology correlation. METHODS: Renal adenocarcinoma cells were injected into the hindlimb of 13 BalB/c mice resulting in tumors ranging from 9.8 to 194.3mm3. US guidance was used to place a laser fiber into the tumor, and VTP was performed. CEUS was performed prior to animal sacrifice, 24h post-VTP. Whole tumors were extracted for histopathologic analysis using H&E and TUNEL staining. Pathology samples corresponding to the CEUS imaging plane were prepared in order to compare the size and extents of tumor necrosis. RESULTS: Tumor necrosis following VTP appeared as a central region of non-enhancement on CEUS, while viable tumor appeared as patchy regions of enhancement in the tumor periphery. The region of tumor necrosis measured in mean 66% and 64.8% of total tumor area on CEUS and pathology respectively (p=0.2). The size and location of the necrosis on CEUS images and pathology samples were found correlative with no inter-observer difference (weighted kappa of 0.771 and 0.823, respectively). CONCLUSION: CEUS allows accurate monitoring of VTP induced tumor necrosis in a small animal model.

Invasion of hepatocytes by Plasmodium sporozoites requires cGMP-dependent protein kinase and calcium dependent protein kinase 4.

Invasion of hepatocytes by sporozoites is essential for Plasmodium to initiate infection of the mammalian host. The parasite's subsequent intracellular differentiation in the liver is the first developmental step of its mammalian cycle. Despite their biological significance, surprisingly little is known of the signalling pathways required for sporozoite invasion. We report that sporozoite invasion of hepatocytes requires signalling through two second-messengers - cGMP mediated by the parasite's cGMP-dependent protein kinase (PKG), and Ca2+ , mediated by the parasite's calcium-dependent protein kinase 4 (CDPK4). Sporozoites expressing a mutated form of Plasmodium berghei PKG or carrying a deletion of the CDPK4 gene are defective in invasion of hepatocytes. Using specific and potent inhibitors of Plasmodium PKG and CDPK4, we demonstrate that PKG and CDPK4 are required for sporozoite motility, and that PKG regulates the secretion of TRAP, an adhesin that is essential for motility. Chemical inhibition of PKG decreases parasite egress from hepatocytes by inhibiting either the formation or release of merosomes. In contrast, genetic inhibition of CDPK4 does not significantly decrease the number of merosomes. By revealing the requirement for PKG and CDPK4 in Plasmodium sporozoite invasion, our work enables a better understanding of kinase pathways that act in different Plasmodium stages.