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The sensitivity of amide, amine, creatine and guanidinium CEST in detecting pH at high MRI field

Lin Chen^1,2 and Jiadi Xu^1,2
¹Department of Radiology and Radiological Science, Johns Hopkins University, Baltimore, MD, United States, ²F.M. Kirby Research Center for Functional Brain Imaging, Kennedy Krieger Research Institute, Baltimore, MD, United States

Synopsis

We compared the sensitivity of several chemical exchange saturation transfer (CEST) contrasts in detecting altered pH at 11.7 T MRI. Studies with egg white phantom revealed that amide and guanidium CEST contrasts in protein are suitable for pH mapping at the physiological relevant range (6.5-7.5), while amineCEST works well for pH lower than 6.5. Hypercapnia (20%) study in mouse brain indicated that creatineCEST showed both higher pH sensitivity and signal intensity compared with amide and guanidium CEST at the physiological pH range and is more suitable for in vivo pH studies at high field.

Purpose

One of the most important applications of CEST MRI is pH mapping, which has already shown great success in many disease diagnoses such as stroke (1), tumor(2), and renal pH homeostasis (3). Up to now, there are many CEST contrasts were discovered for pH mapping at high MRI fields such as amide (3.5 ppm) (1), amine (2.7 ppm) (4), guanidinium (2 ppm) (5), and creatine (2 ppm) (6) CEST. However, there are few comparisons in terms of pH sensitivity among those contrasts. In the current study, we used hypercapnia to alternating the pH in the brain cells (7-10) together with egg white phantom to determine which CEST contrast is suitable for detecting the pH response at the physiological pH range.

Methods

Egg white is an ideal model to demonstrate the sensitivity of the amide, guanidinium, and amine CEST from mobile proteins for detecting pH variation by titrating to pH 6, 6.5, 7, and 7.5. Hypercapnia was used to evaluate the sensitivity of CEST contrast in detecting pH change in brain. Z-spectra on five wild type mouse brains pre- and post-20% CO₂ inhalation were recorded. The 20% CO₂ delivery was accomplished by mixing the air and CO₂ with flow rates of 2 and 0.5 L/min, respectively. CEST experiments were performed using continuous-wave CEST (cwCEST). MR images were acquired using a Turbo Spin Echo (TSE) sequence with TE = 18 ms, TR=5 s, TSE factor = 20, slice thickness =1.5 mm and a matrix size of 64×64. All MRI experiments were performed on a horizontal bore 11.7 T Bruker Biospec system. The saturation field strength (B₁) and length were 2 μT and 1 s for creatine CEST (CrCEST), 0.5 μT and 3 s for guanidinium CEST (guanCEST), and 1 μT and 2 s for amideCEST according to previous optimization results (11-13). The extraction of the CEST signal was achieved using polynomial and Lorentzian line-shape fitting (PLOF) as detailed previously (11,12,14-16).
³¹P MRS was performed to measure the cerebral pH and phosphocreatine changes after CO₂ inhalation. The intracellular pH was calculated from the chemical shift of the Pi peak relative to the PCr peak. The in vivo ³¹P MRS experiments were performed using a single pulse sequence (TR = 2 s, NA = 512, bandwidth = 50 ppm, acquisition time = 100 ms). The total experimental time was 16 minutes.

Results and Discussion

The Z-spectra of egg white phantoms with different pH values are plotted in Fig. 2. The amide peak at 3.5 ppm disappears at pH 6.5 (17). The guanidinium peak increases when pH drops from 7.5 to 6.5. However, the peak will begin to decrease when pH further drops to 6. The amine peak is not visible when pH is higher than 6.5 and will form a single peak centered at 2.7 ppm when the pH is less than 6.5. Interestingly, there still a strong broad signal between 0 to 5 ppm that is not sensitive to pH and is still visible even at pH 6, which is assigned as amideNOE. The amideNOE may be attributable to non-exchanging amide protons or amide proton with extremely slow exchange rates, but they will still be able to transfer magnetization to water following a two-step relayed NOE process with faster exchanging neighboring protons (18). To validate the feasibility of different CEST contrasts (amideCEST, CrCEST and GuanCEST) in detecting subtle cerebral pH changes, Z-spectra were recorded of mouse brain before and after 20% CO2 inhalation (Fig. 2). The results reveal that CrCEST (4.26±0.32%) yields a much stronger signal compared to both amideCEST (1.97±0.39%,) and guanCEST (1.46±0.30%). Both CrCEST and amideCEST exhibited reduced CEST contrast after CO2 inhalation, while guanCEST increases slightly. A significant difference was observed in the change in CrCEST signal intensity (0.90±0.4%, p < 0.001), while a much smaller difference was observed in the amideCEST results (0.59±0.39%, p = 0.061) as well as the guanCEST (-0.07±0.05%, p = 0.74). This observation also confirmed the previous assignment that the CEST peak at 2 ppm is dominated by guanCEST with low saturation powers (<0.6 mT) and is mainly from CrCEST with high powers (11,12).
A cerebral pH change from 7.26±0.07 to 6.99 ±0.07 was measured with 31P MRS. The PCr peak broadened and also the concentration decreased by approximately 9.6±5%, as determined from the integral (2.18 ±1.1 104 to 1.97 ±1.0 104), which is caused by the conversion of PCr to Cr as pH decreases (19,20).
The pH detection sensitivity of CEST MRI is proportional to the change in the exchange rate induced by pH variations. Compared to amide protons, Cr protons possess a much higher exchange rate more than 1000 Hz (21), and hence, a larger exchange rate change related to pH variations.

Conclusion

We demonstrated that CrCEST is a highly pH-sensitive method at high field

Acknowledgements

This work was supported by NIH: R01HL149742, R03NS109664, R21NS118079, R21AG065794 and DOD W81XWH-18-1-0797

References

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Figures

Figure 1: (a) The typical Z-spectra of egg white that titrated to 6.0, 6.5, 7 and 7.5 pH. The spectrum was recorded using CW-CEST with 1μT and 3s saturation at 37°C by an air heater. The CEST peaks from amide, amine, guanidinium, amideNOE are indicated. The simulated DS spectrum for egg white with pH 7.5 is also plotted.

Figure 2. Representative Z-spectra for the GuanCEST experiments (a) pre and (b) post-CO2 inhalation for the cortex region. The PCr peak is observable and is indicated. Representative Z-spectrum of amideCEST experiment (c) pre and (d) post-CO2 inhalation and CrCEST experiment (e) pre and (f) post-CO2 inhalation for the cortex region. Solid lines are the fitted background using the PLOF method. (g) Scatter plots showing the CEST contrast difference for CrCEST, amideCEST and GuanCEST experiments obtained by PLOF (n= 5) and the corresponding contrast changes (h).

Figure 3. (a) Averaged 31P spectra (n=4) for the WT mice pre (red) and post (green) CO2 inhalation. The shift of the inorganic phosphate (Pi) due to the pH change is indicated. (b) Averaged 31P spectra (n=4) post CO2 inhalation for comparing the PCr intensity. The The PCr peaks clearly showed signal reduction and broadening after CO2 inhalation, while the adenosine triphosphate concentration showed a negligible change. A typical T2w image of the mouse brain was plotted to show the position and coverage of the 31P/1H surface coil.

Proc. Intl. Soc. Mag. Reson. Med. 29 (2021)

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