Differentiating Between Cancer and Inflammation: A Metabolic-Based Method for Functional Computed Tomography Imaging

One of the main limitations of the highly used cancer imaging technique, PET-CT, is its inability to distinguish between cancerous lesions and post treatment inflammatory conditions. The reason for this lack of specificity is that [¹⁸F]­FDG-PET is based on increased glucose metabolic activity, which characterizes both cancerous tissues and inflammatory cells. To overcome this limitation, we developed a nanoparticle-based approach, utilizing glucose-functionalized gold nanoparticles (GF-GNPs) as a metabolically targeted CT contrast agent. Our approach demonstrates specific tumor targeting and has successfully distinguished between cancer and inflammatory processes in a combined tumor-inflammation mouse model, due to dissimilarities in angiogenesis occurring under different pathologic conditions. This study provides a set of capabilities in cancer detection, staging and follow-up, and can be applicable to a wide range of cancers that exhibit high metabolic activity

Muscle strength is impaired in rpS6^P−/− mice.

<p>(A to C) Age-matched (4 and 7 month) male mice (n = 13 for WT and 12 for rpS6^P−/− mice) were subjected to screening, according to the SHIRPA behavioral protocol (see details in “Experimental Procedures”). (A) Grip strength. Mice were allowed to grip a grid and a gentle horizontal backwards pull through their tail was applied. Higher scores indicate greater grip strength. An unbiased observer, blinded to the genotype, performed the experiment in a blind fashion; (B) Wire maneuver. Results represent time in seconds required for a mouse that is hung from a wire with its forearms to elevate its hind limbs and grip the wire. (C) Rota-rod performance. Mice were placed on a moving cylinder, which was gradually accelerated from an initial speed of 4 rpm to a maximum of 40 rpm. Latency to fall from the rota-rod is presented in seconds. Motor performance was measured in three 10 min sessions (time 0, 1 h and 24 h). In each trial, the time in seconds until falling off was recorded. *<i>P</i><0.0005 for each trial versus rpS6^P+/+ mice. (D) Endurance test. 5 rpS6^p+/+ and 4 S6^P−/− age-matched (7–9 weeks) male mice were allowed to run on the treadmill set with a slope of 12.5 degree and a speed of 20 m/min. The results represent the total running time with two attempts to pause. Results of all experiments are presented as average±SEM, and the absence of SEM bars for some measurements simply reflects a value close to zero that is graphically invisible.</p

Insulin-induced signaling and glucose uptake are similar in rpS6^P−/− and rpS6^p+/+ muscles.

<p>(A) Soleus muscles were excised from rpS6^P−/− (−/−) and rpS6^p+/+ (WT) 2- to 3-mo-old male mice following 16 h starvation and intraperitoneal injection of saline (−) or 2.5 U of insulin/kg of body weight (+) for 5 min. Cytoplasmic extracts were subjected to Western blot analysis with the indicated antibodies. Note the enrichment for the upper band of 4E-BP1 (hyperphosphorylated form) upon insulin treatment. (B) Insulin-induced uptake of glucose into isolated muscle. Right and left soleus muscles were isolated from the hind limbs of 16 h starved rpS6^P−/− (−/−) and rpS6^p+/+ (WT) male mice. 2-Deoxy glucose uptake is presented as µmol per gram tissue per h. The data are presented as average±SEM for 5 rpS6^P−/− and 4 S6^p+/+ age-matched (7–9 weeks) male mice. (C) Mice were injected intraperitoneally with 0.25 U insulin/kg body weight and within seconds with 10–24 Ci [¹⁸F]fluoro-2-deoxyglucose (FDG) into the tail vein. The radioactivity concentration in hind limb muscles was estimated at 40 to 45 min after FDG injection by positron emission tomography. The concentration of radioactivity is presented as % injected dose (ID) per gm body weight per ml tissue. The data are shown as average±SEM for 5 male mice of each genotype.</p

The amount of mitochondria and the activity of complexes of the oxidative phosphorylation system are unchanged in rpS6^P−/− muscle.

<p>Mitochondria were isolated from hind limb muscle of four age-matched rpS6^P+/+ (WT) and rpS6^P−/− (−/−) male mice and were assayed for citrate synthase (A), cytochrome C oxidase (B), as well as complexes I (C), II (D), II+III (E) and IV (F) of the oxidative phosphorylation system. The activity of the different complexes was normalized to that of citrate synthase and the results are presented as relative activities. Vertical bars represent SEM (n = 4 age- matched male mice for each genotype).</p

Newly formed myofibers are smaller in a rpS6^P−/− muscle.

<p>(A) Sections of tibialis anterior of 2-mo-old rpS6^P+/+ and rpS6^P−/− mice were prepared 5 days after cryoinjury, and were immunostained for eMHC. One representative section is presented. (B) Enlargement of the framed area in (A). (C) Digital images of the immunostained sections were used for measurement of CSA of individual eMHC-positive myofibers, as described in “Experimental Procedures”. The average CSA of rpS6^P−/− myofibers (n = 327) was normalized to that of their wild type counterparts (n = 210), which was arbitrarily set at 1. The results are presented as average±SEM. *, p<0.0001.</p

Stores of glycogen and triacylglycerol are larger and similar, respectively, in rpS6^P−/− muscle.

<p>(A) Soleus glycogen content. Soleus muscles were excised from 20 rpS6^P+/+ (WT) and 15 age-matched rpS6^P−/− (−/−) male mice and their glycogen content was measured. (B) Glycogen synthase (GS) and (C) glycogen phosphorylase (GP) activities were assayed in the same extracts (n = 4 age-matched male mice for each genotype). (D) and (E) TG content in soleus and gastrocnemious, respectively. 5 soleus (left and right muscles were pooled) and 10 individual gastrocnemious muscles were excised from 5 rpS6^P+/+ (WT) and 5 age-matched rpS6^P−/− (−/−) male mice and their TG content was measured. All results are presented as average±SEM.</p