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Abstract

Background. [¹⁸F]FEPPA is a potent TSPO imaging agent that has been found to be a potential tracer for imaging neuroinflammation. In order to fulfill the demand of this tracer for preclinical and clinical studies, we have developed a one-pot automated synthesis with simplified HPLC purification of this tracer, which was then used for PET imaging of neuroinflammation in fine particulate matter- (PM2.5-) exposed rats. Results. Using this automated synthesis method, the RCY of the [¹⁸F]FEPPA was $38 \pm 4 %$ ( $n = 17$ , EOB) in a synthesis time of $83 \pm 8$ min from EOB. The radiochemical purity and molar activities were greater than 99% and $209 \pm 138$ GBq/μmol (EOS, $n = 15$ ), respectively. The quality of the [¹⁸F]FEPPA synthesized by this method met the U.S. Pharmacopoeia (USP) criteria. The stability test showed that the [¹⁸F]FEPPA was stable at $21 \pm 2$ °C for up to 4 hr after the end of synthesis (EOS). Moreover, microPET imaging showed that increased tracer activity of [¹⁸F]FEPPA in the brain of PM2.5-exposed rats ( $n = 6$ ) were higher than that of normal controls ( $n = 6$ ) and regional-specific. Conclusions. Using the improved semipreparative HPLC purification, [¹⁸F]FEPPA has been produced in high quantity, high quality, and high reproducibility and, for the first time, used for PET imaging the effects of PM2.5 in the rat brain. It is ready to be used for imaging inflammation in various clinical or preclinical studies, especially for nearby PET centers without cyclotrons.

1. Introduction

Neuroinflammation is an inflammatory and adaptive response within the central nervous system (CNS) [1] and is the driving force for disease progression, such as Alzheimer’s disease (AD) [2], major depressive disorder [3], schizophrenia [4], and brain injuries [4]. Activated in response to neuropathologies [5, 6], microglia and astrocytes have been known to be the predominant mediators during the neuroinflammatory process. The transmembrane domain protein, translocator protein-18 kDa (TSPO), is variously expressed throughout the body and has low expression within the brain [7–9]. However, TSPO levels are significantly increased in the brain when microglia and astrocyte are activated [10, 11]. Recently, the results of meta-analyses of TSPO levels in mild cognitive impairment and AD further supported the association of increased neuroinflammation during the progression of mild cognitive impairment and AD, relative to healthy controls [12]. In addition to CNS, an increase in TSPO expression has also been seen in a wide variety of malignant human cells and tissues including brain cancers [13, 14], prostate cancers [15, 16], colon cancers [17–19], breast cancers [20, 21], esophageal cancers [22], endometrial carcinomas [23], ovarian cancers, and hepatic carcinomas [24]. Albeit its lack of specificity to activated microglia, the TSPO levels may at times reflect neuroinflammation in vivo [10].

Exposure to fine particulate matter (PM2.5) has been linked to adverse neurological and behavioral health effects, including increased risk for cognitive decline (AD) [25, 26], Parkinson’s disease [27], ischemic stroke [28], and anxiety or depression [29, 30] in epidemiology studies. Increases in neuroinflammation and oxidative stress have been identified as putative mechanisms by which fine PM2.5 may impair central nervous system function [31, 32], an example of which occurs when exacerbated and unregulated microglial proinflammatory responses play crucial factors involved in brain damage [33]. Chronic activation of the microglia (reactive microgliosis) has been implicated in neuronal injury and neuronal damage after exposure to PM2.5 [34]. One hallmark of microglial activation is the overexpression of TSPO [35, 36]. To the best of our knowledge, the application of a TSPO imaging agent for studying the effects of PM2.5 on human health has not been reported. Thus, we have developed an one-pot automated synthesis of [¹⁸F]FEPPA, a potent TSPO imaging agent, and used it to investigate (1) whether PET imaging can noninvasively detect microglia activation after chronic subacute ambient PM2.5 exposure in spontaneous hypertensive rats and (2) whether there is a specific pattern of microglia activation in the brain after chronic subacute ambient PM2.5 exposure.

The first TSPO PET ligand, [¹¹C]PK-11195 was synthesized more than two decades ago [37–39]. However, its utility was limited due to its low brain penetration, high nonspecific binding, high plasma protein binding, and short half-life (for review, see [37, 38]). Therefore, several second-generation ¹⁸F-labelled TSPO PET ligands, including [¹⁸F]FEPPA [40–43], [¹⁸F]FEDAA1106 [44, 45], [¹⁸F]DPA-714 [46, 47], and [¹⁸F]PBR06 [48, 49] have been developed (for review, see [50, 51]). Recently, a novel third-generation TSPO PET ligand, [¹⁸F]GE180 was developed [52–54] and proved to be useful for TSPO imaging [53, 55, 56]. However, its application for neuroimaging is very limited as it suffers from very low brain penetration, similar to that of [¹¹C]PK-11195. Specifically, in humans, the VT of [¹⁸F]GE180 is 20-fold lower than that of [¹¹C]PRB28 [57]. Among these tracers, [¹⁸F]-N-(2-(2-fluoroethoxy)benzyl)-N-(4-phenoxypyridin-3-yl)acetamide ([¹⁸F]FEPPA) shows outstanding properties regarding affinity, stability, lipophilicity, and radiosynthesis [43] and has been used in several preclinical [40, 58, 59] and clinical settings [60–66]. In order to facilitate the usefulness of [¹⁸F]FEPPA in both preclinical and clinical studies, it is imperative to have a fully automated, simple, high yield, and reliable manufacturing process method available for the production of this tracer. Thus, we have adopted Wilson’s method [43], with some modifications, to fully automate the synthesis of this potent TSPO imaging agent using a TRACERlab Fx_FN module (GE Healthcare, Milwaukee, WI) with ethanol and water for purification in high quality and high reproducibility. Vignal et al. reported a similar synthesis using ethanol, water, and phosphoric acid for purification of [¹⁸F]FEPPA [42]. We report herein (1) a one-pot automated synthesis of [¹⁸F]FEPPA with a simplified HPLC purification, (2) the USP compliant QC and stability tests of [¹⁸F]FEPPA, and (3) microPET imaging of [¹⁸F]FEPPA in PM2.5-exposed rats.

2. Materials and Methods

The precursor (N-[[2-[2-[[(4-methylphenyl)sulfonyl]oxy]ethoxy]phenyl]methyl]-N-(4-phenoxy-3-pyridinyl) acetamide, TsEPPA, 1) and the nonradioactive authentic sample N-(2-(2-fluoroethoxy)benzyl)-N-(4-phenoxypyridin-3-yl) acetamide (FEPPA, 2) were purchased from ABX Advanced Biochemicals (Radeberg, Germany). All other chemicals and solvents were purchased from either Sigma-Aldrich (Milwaukee, WI, USA) or Acros Organics (Morris Plains, NJ, USA) and used without further purification. [¹⁸O]O₂H (>98% enriched) was purchased from Rotem Industries (Beer Sheva, Israel). Aqueous [¹⁸F]Fluoride was produced in our PET Trace cyclotron (GE Medical Systems, Uppsala, Sweden) via ¹⁸O(p, n)¹⁸F nuclear reaction. All Sep-Pak® cartridges were purchased from Waters Associates (Milford, MA, USA).

2.1. Automated Radiosynthesis

The [¹⁸F]FEPPA (2) was produced by automated synthesis via fluorination of the TsEPPA (1) with K[¹⁸F]/K_2.2.2 followed by purification with HPLC, to give [¹⁸F]FEPPA (2) as previously reported (Scheme 1) [43].

Scheme 1

Radiosynthesis of the [¹⁸F]FEPPA (2).

The [¹⁸F]FEPPA (2) was produced by automated synthesis using a modified TRACERlab Fx_FN module (GE Healthcare, Milwaukee, WI; Figure 1). Briefly, fluorination of TsEPPA (1) with K[¹⁸F]/K_2.2.2 in anhydrous MeCN at 70°C for 20 min gave the crude product (1) (Scheme 1). After dilution with H₂O, the crude product (1) was purified with a semipreparative HPLC (Waters Xterra RP-18, 10 μm, $10 \times 250$ mm, 35% aqueous ethanol in water for injection, 254 nm, 4 mL/min). The fraction containing [¹⁸F]FEPPA (2) was collected, reformulation, and sterile filtration to provide (2) with <10% of ethanol concentration for PET imaging of the effects for PM2.5 in rats. Please refer to Supplementary data for detailed steps of radiosynthesis.

Figure 1

Modified TRACERLab Fx_FN module for the [¹⁸F]FEPPA (2) radiosynthesis.

2.2. Quality Control (QC) and Stability Tests of the [¹⁸F]FEPPA (2)

The radiochemical purity, chemical purity, and molar activity of 2 were analyzed with an HPLC system (Agilent 1100 series) equipped with a Bioscan FC3300 flow count radioactivity detector ( $2^{″} \times 2^{″}$ pinhole) and a UV detector (254 nm) using a Waters Xterra column (RP-18, 5 μm, $4.6 \times 250$ mm) with 50% aqueous MeCN as the eluent and a flow rate of 1.0 mL/min. The chemical identity of the [¹⁸F]FEPPA (2) was confirmed by coinjection with a nonradioactive authentic FEPPA. The radiochemical impurity of 2 was further analyzed with radio TLC (Silica gel 60 F254 plate, 10 cm, ethyl acetate/hexane (3/1)) and detected with a Raytest miniGita radio-TLC scanner (Raytest, Straubenhardt, Germany). Please refer to Supplementary data and Figure S1 for details.

Other items of QC test and corresponding criteria of 2 were set according to the U.S. Pharmacopoeia (USP) for radiopharmaceuticals [67], which included visual inspection, pH, half-life of radionuclide, radionuclidic purity, radiochemical purity, chemical purity, residual K_2.2.2, residual solvents, bacterial endotoxins, filter integrity, and sterility test.

The stability of 2 at $21 \pm 2$ °Cwas monitored with both TLC and HPLC as described above for up to 4 hr after EOS.

2.3. MicroPET Imaging of the [¹⁸F]FEPPA (2) Injection in Rats Exposed to Ambient Fine Particulate Matter (PM2.5)

2.3.1. Animals

Male spontaneously hypertensive rats (7-week-old, average weight of 350 g, $n = 12$ ) were obtained from the National Laboratory Animal Center (Taipei, Taiwan) and were randomly classified into ambient fine PM exposure group (aerodynamic diameter of <2.5 μm, PM2.5; $n = 6$ ) and high-efficiency particulate air- (HEPA-)-filtered air (FL, $n = 6$ ) conditions group using Taipei Air Pollution Exposure System (TAPES) for health effects [68, 69] for 24 hours per day, 7 days per week, for a total of 6 months. The coarse (PM2.5-10) and fine (PM2.5) size concentrations constituted 0.4 and 99.6% of the whole-body exposure system [70]. Therefore, the rats were mostly exposed to PM2.5. The rats were housed in ventilated cages under a conditioned environment as illustrated in our previous studies [68]. Lab diet and water were provided ad libitum during the study. The animal experiments adhered to protocols of the Institutional Animal Care and Use Committee (IACUC) of the Laboratory Animal Center at National Taiwan University (ACUC no: 20160025).

At the end of exposure, [¹⁸F]FEPPA microPET/CT brain scan was performed for each rat to assess the degree of neuroinflammation. During scanning, rats were anesthetized by passive inhalation of a mixture of isoflurane (5% for induction and 2% for maintenance) in oxygen, followed by a bolus tail vein injection of $24 \pm 10$ MBq, $n = 6$ ) of (2). Brain transmission and emission scans of rats were acquired with a small animal Argus PET/CT scanner (SEDECAL, Madrid, Spain). Ten minutes posttail vein injection of (2), static sinograms were produced for 30 min for each organ of interest. Images were analyzed using commercial PMOD software (PMOD Technologies LLC, version 3.6, Zurich, Switzerland) after coregistration the microCT images to the innate MR atlas. Regional brain radioactivity concentrations (MBq/cm³) were normalized to the injected activity (MBq) and the weight (g) of organ to yield a semiquantitative measurement of radiotracer binding, standardized uptake value (SUV) [71]. The mean SUV of each brain region (volume of interest, VOI) such as the whole brain, frontal lobe, parietal lobe, insular lobe, occipital lobe, midbrain, temporal lobe, hippocampus, retrosplenial cortex in the temporal lobes, and cerebellum were calculated and analyzed.

2.4. Immunohistochemical Analysis

After PET image acquisition, all rats were sacrificed. The brain tissues were processed using an automated tissue processor (Shandon Excelsior, Thermo Scientific, UK), embedded in paraffin, and cut at a thickness of 3-5 μm for immunohistochemical (IHC) staining. Ionized calcium binding adaptor molecule (Iba1) is a macrophage-specific calcium-binding protein, participating in membrane ruffling and phagocytosis in the activated microglia. For Iba1 staining, brain tissues were stained against the activated microglial marker Iba1 (GeneTex, San Antonio, TX, USA) [72]. The percentage of areas occupied by the stained nuclei of activated microglia at 40x high power field (0.26 mm/pixel) in the hippocampal region of the rats was calculated using a digital slide scanner (MoticEasyScan, Motic®, Canada). The brain tissues were examined by a histopathologist blinded to the exposure data.

2.5. Statistical Analysis

Data was expressed as $mean \pm standard deviation (SD)$ . Wilcoxon rank sum test was used to compare different regions of brain in PM2.5 and FL rats. $P < 0.05$ was regarded as statistically significant. Statistical analyses were performed using JMP®, version 10 statistical software package (SAS Institute Inc., Cary, NC, USA).

3. Results

3.1. A One-Pot Automated Synthesis of the [¹⁸F]FEPPA (2) Using a Modified Fx_FN Module

In this paper, we presented the detailed automated production of [¹⁸F]FEPPA along with a full set of quality control specifications and results in USP compliance. Using this modified purification method, we were able to routinely produce 2 with a Fx_FN module in $38 \pm 4 %$ yield (EOB, $n = 17$ ) in a synthesis time of $83 \pm 8$ min from EOB. Typically, starting with 35 GBq of [¹⁸F]Fluoride, 8 GBq of 2 was produced at EOS. Both the chemical and radiochemical purities of 2 were greater than 90% with a molar activity of $209 \pm 138$ GBq/μmol ( $n = 15$ , EOS) and were used for PET imaging of the effects of PM2.5 in rats.

The QC test results of 2 are tabulated in Table 1. Moreover, the stability test showed that 2 synthesized by this method was stable at room temperature ( $21 \pm 2$ °C) for up to 4 hr after EOS (Table 2). Detailed QC data for [¹⁸F]FEPPA produced using our method disclosed herein are shown in supplementary data.

Table 1

The QC tests of the [¹⁸F]FEPPA (2).

Items tested	Acceptance criteria
Visual inspection	Clear, colorless solution	Pass	Pass	Pass
pH value	5-8	7	7	7
Residual K_2.2.2 (μg/mL)	<50	<50	<50	<50
Radionuclidic purity (%)	>99.5	99.93	99.96	99.62
Half-life (min)	$110 \pm 5$	108	115	115
Radiochemical identity (min)	∣R_t-R_t (reference)∣≤0.5	0.02	0.00	0.07
Radiochemical purity (%)	>90	99.9	99.8	99.8
Residual solvent analysis	$EtOH < 10 %$ $Acetone < 0.5 %$ $MeCN < 0.04 %$	4.6849% 0.1454% 0.0100%	0.1673% 0.0324% 0.0014%	2.2298% 0.0010% 0.0008%
Residual [¹⁸F]Fluoride (%)	<5	0.01	0.02	0.59
Bacterial endotoxins (EU/V)	<175	<17.5	<17.5	<17.5
Filter integrity test (psi)	>45	48.5	46.6	46.0
Sterility test	Sterile	Sterile	Sterile	Sterile

Table 2

Stability of three consecutive productions of the [¹⁸F]FEPPA (2) ( $n = 3$ ).

Items tested	Acceptance criteria	Elapsed time (hr)	Run 1	Run 2	Run 3
Radiochemical purity (%)	>90%	0	99.9	99.8	99.8
		2	99.0	99.3	98.6
		4	98.9	99.4	97.7
Residual [¹⁸F]Fluoride (%)	<5%	0	0.01	0.02	0.59
		2	0.34	0.73	1.01
		4	1.04	0.23	1.89

3.2. MicroPET Imaging of the [¹⁸F]FEPPA (2) Injection in Rats Exposed to Ambient Fine Particulate Matter (PM2.5)

The mean mass concentration of PM2.5 during the exposure period was $10.8 \pm 3.8$ μg/m³. The predominant chemical composition of the trapped particles in the PM group was sulfur (16.0% of mean PM2.5 concentration), potassium (1.2%), and iron (0.6%), similar to that in previous PM exposure data [68, 70]. There was no significant alterations in the weight of the rats after ambient PM2.5 exposure between the PM2.5 and FL groups.

Typical whole-brain biodistribution of [¹⁸F]FEPPA in the PM2.5 and FL rats was depicted in Figure 2. Increased tracer activity of [¹⁸F]FEPPA in selected brain regions was shown in Table 3 and Figure 3 and expressed as SUV mean.

Figure 2

Representative imaging of coronal, sagittal, and axial sections of rat brain exposed to ambient PM2.5 ((a) PM2.5, left panel) versus filtered air ((b) FL, right panel).

(b)

Table 3

MicroPET Imaging of the [¹⁸F]FEPPA (2) in ambient PM2.5-exposed ( $n = 6$ ) versus filtered air-exposed ( $n = 6$ ) rats.

Region	PM (SUV mean)	FA (SUV mean)	$P$ value
Whole brain	$0.85 \pm 0.02$	$0.78 \pm 0.03$	0.03 $^{*}$
Frontal lobe	$0.92 \pm 0.02$	$0.87 \pm 0.02$	0.047
Parietal lobe	$0.73 \pm 0.04$	$0.75 \pm 0.03$	NS
Insular lobe	$0.93 \pm 0.04$	$0.83 \pm 0.02$	0.047
Occipital lobe	$0.64 \pm 0.04$	$0.68 \pm 0.03$	NS
Midbrain	$0.80 \pm 0.04$	$0.81 \pm 0.04$	NS
Olfactory bulb	$0.99 \pm 0.04$	$1.06 \pm 0.06$	NS
Temporal lobe	$0.78 \pm 0.02$	$0.71 \pm 0.03$	0.04 $^{*}$
Hippocampus	$0.73 \pm 0.02$	$0.66 \pm 0.02$	0.01 $^{*}$
Retrosplenial cortex	$0.83 \pm 0.04$	$0.71 \pm 0.04$	0.03 $^{*}$
Cerebellum	$0.85 \pm 0.04$	$0.86 \pm 0.05$	NS

Abbreviations: FA: filtered air; NS: not significant; PM: particulate matter.

Figure 3

Tracer distribution of the [¹⁸F]FEPPA (2) in the hippocampus (a) and retrosplenial region (b) of the temporal lobes.

(b)

Specifically, significant increased [¹⁸F]FEPPA tracer activity was observed in the temporal lobe of the PM2.5 compared to that of the FL rats ( $P = 0.04$ ), especially in the hippocampus (Figure 2, red contour, $P = 0.01$ ) and retrosplenic region (Figure 2, green contour, $P = 0.03$ ). A trend of increased tracer activity was found at insular areas (Figure 2, indigo contour) and frontal lobe (not shown) in the PM2.5 rats, but not reaching statistical difference (Table 3).

3.3. Immunohistochemical (IHC) Staining of the Rat Brain

The IHC staining of the hippocampus showed that the % of area with Iba1 staining at 40x high power field were $0.130 \pm 0.017 %$ in the FL group vs. $0.313 \pm 0.081 %$ in the PM2.5 group (Figure 4, $P = 0.05$ ).

Figure 4

The immunostaining (brown-colored nuclei, red arrows) in the hippocampus of the PM rats (a), FL rats (b), and the differences between these two groups (c).

(b)

(c)

There is a trend for higher Iba1 staining (brown-colored nuclei, red arrows) in the hippocampus of PM rats (a) compared to the FL group (b), but the difference is not statistically significant (staining area of $0.313 \pm 0.081 %$ vs. $0.130 \pm 0.017 %$ , $P = 0.05$ (c)).

4. Discussion

[¹⁸F]FEPPA is a potent TSPO imaging agent that has been found to be a potential tracer for imaging neuroinflammation. In order to fulfill the demand of this tracer for preclinical and clinical studies, we have developed a one-pot automated synthesis with simplified HPLC purification of this tracer, which was then, for the first time used for PET imaging of the effects of PM2.5 in rats.

[¹⁸ F]FEPPA (2) has been synthesized by several methods in various radiochemical yields and radiochemical purities [42, 43, 58, 73]. Initially, 2 was synthesized manually by nucleophilic fluorination of the corresponding tosylate-precursor (TsEPPA, 1) with [¹⁸F]Fluoride, followed by purification with HPLC, to give 2 in 71~85% yield (EOB) (Scheme 1) [43]. Later, several automated syntheses of 2 were reported [42, 58, 73] (Table 4).

Table 4

Synthesis of the [¹⁸F]FEPPA (2) via various methods.

References	Radiofluorination condition				RCY (%, EOB)	Synthesis time (min)	Number of production	Molar activity (GBq/μmol)	Automate synthesizer	QC (EP or USP)	TLC
References	Precursor weight (mg)	Solvent (mL)	Temperature (°C)	Time (min)	RCY (%, EOB)	Synthesis time (min)	Number of production	Molar activity (GBq/μmol)	Automate synthesizer	QC (EP or USP)	TLC
Wilson et al. [43]	5	MeCN 0.7 mL	90	10	71–85	40–50	—	44~100	—	—	—
Vasdev et al. [58]	5	MeCN 0.75 mL	90	10	$38 \pm 10$	$36 \pm 1$	54	$222 \pm 148$	Fx_FN	—	—
Berroterán-Infante et al. [73]	3.125	MeCN 0.5 mL	90	10	$38 \pm 3$	30	15	$241 \pm 13$	Nuclear Interface	EP	Y $^{*}$
Vignal et al. [42]	5	MeCN 1 mL	90	10	$48 \pm 3$	55	17	$198 \pm 125$	AllInOne®	—	—
Zammit et al. [59]	2	MeCN 0.4 mL	90	10	$41 \pm 8$	$78 \pm 4$	—	$751 \pm 163$	CPCU	—	—
Chang et al. [74]	5	MeCN 0.6 mL	90	10	50	80	8	149~217	Eckert-Ziegler modular system	—	—
This study	5	MeCN 1 mL	70	20	$38 \pm 4$	$83 \pm 8$	17	$209 \pm 138$	Fx_FN	USP	Y

$^{*}$ No TLC data were provided.

However, most of these automated syntheses used toxic MeCN as the solvent for semipreparative HPLC purification, and thus, reformulation was necessary. As a result, we and others [42] have chosen to use aqueous ethanol as the mobile phase for purification of 2 with HPLC. The HPLC purification conditions of 2 have been optimized by using a Waters Xterra RP-18 column (10 μm, $10 \times 25$ mm) and eluting with different flow rates and different concentrations of aqueous ethanol as mobile phase (Figure S2). Although the retention time was slightly longer when separation was done using 35% instead of 40% EtOH, but because of practical considerations, including nonradioactive impurity separation and easy-to-prepare mobile phase (Figure S3), 35% EtOH at a flow rate of 4 mL/min was chosen as the best condition for purification 2 (Figure 5).

Figure 5

Representative semipreparative HPLC purification chromatogram of the [¹⁸F]FEPPA (2).

However, a small amount of [¹⁸F]Fluoride was detected in the eluate, which may have had an impact on PET image quality [74]. Thus, the eluate was further passed through an Alumina-N cartridge and a PTFE sterile filter in series to remove the residual [¹⁸F]Fluoride and resulted in the pure and sterile 2 in $38 \pm 4 %$ yield (EOB, $n = 17$ ) in a synthesis time of $83 \pm 8$ min from EOB, which is comparable to 2 synthesized by other methods (Table 4). For TSPO-targeting studies, the molar activity is recommended as high as possible as required for studies targeting receptors. Compared to previous published methods, our method achieved a reasonable and high molar activity of $209 \pm 138$ GBq/μmol, EOS ( $n = 17$ ) as shown in Table 4. Quality control and stability tests confirmed that 2 synthesized by this method met the USP. As far as we know, our method is still the only to address residual [¹⁸F]Fluoride concern during automated production, as well as the QC item of residual [¹⁸F]Fluoride determination using radio-TLC analysis, in order to achieve high quality of produced [¹⁸F]FEPPA for preclinical or clinical studies.

PET imaging in rats showed that the whole-brain [¹⁸F]FEPPA tracer activity of the PM2.5 rats was significantly higher than that of their age-matched FL rats. In addition, the brain tracer activity of [¹⁸F]FEPPA was found to be regional-specific, and the difference was more pronounced in the temporal and insular lobes. In the temporal lobe, only the hippocampus and retrosplenial cortex showed statistical higher [¹⁸F]FEPPA activity in the PM2.5 rats when compared to that of the FL rats (Table 3, Figure 3). A trend of increased tracer activity was found at both insular and frontal regions in the PM2.5 rats but did not reach a statistically significant difference.

The IHC staining results showed that although there was not a statistically significant difference in the hippocampus, a trend of increased Iba1 staining in the microglia cells was observed in the PM2.5 group compared to that of the filtered air group, which suggested that microglial activation and inflammation, especially in the temporal lobe, may play important roles in the response of the brain to traffic-related PM.

Taken collectively, we have developed an improved method to automatically produce [¹⁸F]FEPPA in high quantity, high quality, and high reproducibility, and for the first time using it to noninvasively elucidate the microglial changes in different brain regions of rats after subchronic real-world exposure to ambient PM2.5. Our study confirmed that chronic subacute ambient PM2.5 exposure can lead to diffuse microglial activation. The brain area that was especially vulnerable to PM2.5 effects was the temporal lobe, particularly the hippocampus and retrosplenial cortex. These regions played crucial roles in memory, learning, and navigation, especially in the hippocampus. However, there are several limitations of the present rodent study. First, the limited sample size may influence the overall power of the test. On the other hands, this also suggests that with larger sample sizes, perhaps more brain regions will be affected, as we did observe a trend of increased [¹⁸F]FEPPA activity (not reaching statistical significance) in the insular and frontal regions of PM2.5 rats compared to their controls. Second, hypertensive rats were used in the current experiment, a subgroup that may be more vulnerable to fine PM exposure than normotensive rats. Spontaneous hypertensive rats were chosen since cardiovascular diseases are recognized as risk factors for developing neurodegenerative disease in humans and therefore may be more susceptible to traffic-related PM2.5 exposure. Third, blocking experiment using a nonradioactive FEPPA or other nonradioactive TSPO ligand was not performed in the current study. Nonetheless, [¹⁸F]FEPPA is a relatively popular ligand that had been used in many preclinical and clinical studies [75, 76], including and not limited to psychiatric disorders [62, 77]. Various studies have also demonstrated that [¹⁸F]FEPPA reliably binds to TSPO in the brain [78–80]. Lastly, not all regions of the brain were analyzed for Iba1 staining. Nevertheless, the areas of increased Iba1 staining were consistent with the [¹⁸F]FEPPA PET findings.

5. Conclusions

With this one-pot automated synthesis and improved purification of the [¹⁸F]FEPPA (2) using a Fx_FN Module, [¹⁸F]FEPPA (2) can be produced with high quality, quantity, and reproducibility. PET imaging in rats exposed to low-level PM2.5 demonstrated that pulmonary exposure to fine PM may exert health effects on specific areas of the brain, including the hippocampus. The microglial activation and inflammation can be noninvasively evaluated and followed by [¹⁸F]FEPPA PET imaging. The role of microglial response after low-level fine PM2.5 exposure warrants further investigation in humans. The applications of [¹⁸F]FEPPA (2) for studying inflammation in the peripheral organs of animals and humans are in progress.

Footnotes

Data Availability

The datasets used and/or analyzed during the current study are available from the corresponding authors on reasonable request.

Ethical Approval

The animal experiments adhered to the Guide for the Care and Use of Laboratory Animals and were approved by the Laboratory Animal Center at National Taiwan University (Taipei, Taiwan).

Conflicts of Interest

The authors declare that they have no competing interests.

Authors’ Contributions

MFC, TJC, YLG, HMW, WSH, and CYS were responsible for the study conception and design. TJC, HMW, and WSH were responsible for the grant support. YYH and CHC performed the radiopharmaceutical synthesis and quality control. MFC did the animal experiments. MFC and RFY interpreted the microPET images and performed the data analysis. MFC and YYH were major contributors in writing the manuscript. CYS critically reviewed and edited the manuscript for intellectual content. All authors read and approved the final manuscript. Ya-Yao Huang, Wen-Sheng Huang, and Chyng-Yann Shiue contributed equally to this work.

Acknowledgments

We are grateful to Ms. Kwanyu Lin, Ms. Yu-Ning Cheng, Mr. Pei-Yau Lin, and Mr. Chi-Han Wu, for their technical support. This work was supported by the Ministry of Science and Technology of Taiwan (MOST 103-2314-B-002-040-MY3, MOST 105-2314-B-075-060-MY3, and MOST109-2314-B-002-101) and National Taiwan University Hospital (NTUH 109-S4475).

Supplementary Materials

References

DiSabato

D. J.

Quan

Godbout

J. P.

Neuroinflammation: the devil is in the details

Journal of Neurochemistry 2016 139

1076444

Suppl 2 136 153

10.1111/jnc.13607

2-s2.0-84964928591

26990767

Calsolaro

Edison

Neuroinflammation in Alzheimer's disease: current evidence and future directions

Alzheimers Dement 2016 12

1076444

6 719 732

10.1016/j.jalz.2016.02.010

2-s2.0-84973520116

27179961

Troubat

Barone

Leman

Desmidt

Cressant

Atanasova

Brizard

el Hage

Surget

Belzung

Camus

Neuroinflammation and depression: a review

The European Journal of Neuroscience 2021 53

1076444

1 151 171

10.1111/ejn.14720

Marques

T. R.

Ashok

A. H.

Pillinger

Veronese

Turkheimer

F. E.

Dazzan

Sommer

I. E. C.

Howes

O. D.

Neuroinflammation in schizophrenia: meta-analysis of in vivo microglial imaging studies

Psychological Medicine 2019 49

1076444

13 2186 2196

10.1017/S0033291718003057

2-s2.0-85055487782

30355368

Meda

Cassatella

M. A.

Szendrei

G. I.

Otvos

Jr. Baron

Villalba

Ferrari

Rossi

Activation of microglial cells by β-amyloid protein and interferon-γ

Nature 1995 374

1076444

6523 647 650

10.1038/374647a0

2-s2.0-0028953529

Morales

Jimenez

J. M.

Mancilla

Maccioni

R. B.

Tau oligomers and fibrils induce activation of microglial cells

Journal of Alzheimer's Disease 2013 37

1076444

4 849 856

10.3233/JAD-131843

2-s2.0-84892695369

23948931

Anholt

R. R.

De Souza

E. B.

Oster-Granite

M. L.

Snyder

S. H.

Peripheral-type benzodiazepine receptors: autoradiographic localization in whole-body sections of neonatal rats

The Journal of Pharmacology and Experimental Therapeutics 1985 233

1076444

2 517 526

2987488

Bribes

Carriere

Goubet

Galiegue

Casellas

Simony-Lafontaine

Immunohistochemical assessment of the peripheral benzodiazepine receptor in human tissues

The Journal of Histochemistry and Cytochemistry 2004 52

1076444

1 19 28

10.1177/002215540405200103

2-s2.0-0348048881

14688214

Gehlert

D. R.

Yamamura

H. I.

Wamsley

J. K.

Autoradiographic localization of "peripheral-type" benzodiazepine binding sites in the rat brain, heart and kidney

Naunyn-Schmiedeberg's Archives of Pharmacology 1985 328

1076444

4 454 460

10.1007/BF00692915

2-s2.0-0021949310

2986017

10.

Lavisse

Guillermier

Herard

A. S.

Petit

Delahaye

van Camp

Ben Haim

Lebon

Remy

Dolle

Delzescaux

Bonvento

Hantraye

Escartin

Reactive astrocytes overexpress TSPO and are detected by TSPO positron emission tomography imaging

The Journal of Neuroscience 2012 32

1076444

32 10809 10818

10.1523/JNEUROSCI.1487-12.2012

2-s2.0-84864851281

22875916

11.

Wilms

Claasen

Rohl

Sievers

Deuschl

Lucius

Involvement of benzodiazepine receptors in neuroinflammatory and neurodegenerative diseases: evidence from activated microglial cells in vitro

Neurobiology of Disease 2003 14

1076444

3 417 424

10.1016/j.nbd.2003.07.002

2-s2.0-0348109489

14678758

12.

Bradburn

Murgatroyd

Ray

Neuroinflammation in mild cognitive impairment and Alzheimer's disease: a meta-analysis

Ageing Research Reviews 2019 50

1076444

1 8

10.1016/j.arr.2019.01.002

2-s2.0-85059479257

30610927

13.

Miettinen

Kononen

Haapasalo

Helén

Sallinen

Harjuntausta

Helin

Alho

Expression of peripheral-type benzodiazepine receptor and diazepam binding inhibitor in human astrocytomas: relationship to cell proliferation

Cancer Research 1995 55

1076444

12 2691 2695

7780986

14.

Miyazawa

Hamel

Diksic

Assessment of the peripheral benzodiazepine receptors in human gliomas by two methods

Journal of Neuro-Oncology 1998 38

1076444

1 19 26

10.1023/A:1005933226966

2-s2.0-0031930856

15.

Fafalios

Akhavan

Parwani

A. V.

Bies

R. R.

McHugh

K. J.

Pflug

B. R.

Translocator protein blockade reduces prostate tumor growth

Clinical Cancer Research 2009 15

1076444

19 6177 6184

10.1158/1078-0432.CCR-09-0844

2-s2.0-70349667466

19789311

16.

Han

Slack

R. S.

Papadopoulos

Expression of peripheral benzodiazepine receptor (PBR) in human tumors: relationship to breast, colorectal, and prostate tumor progression

Journal of Receptor and Signal Transduction Research 2003 23

1076444

2-3 225 238

10.1081/RRS-120025210

2-s2.0-0242299669

14626449

17.

Katz

Eitan

Gavish

Increase in peripheral benzodiazepine binding sites in colonic adenocarcinoma

Oncology 2004 47

1076444

139 142

18.

Königsrainer

Vogel

U. F.

Beckert

Sotlar

Coerper

Braun

Lembert

Steurer

Königsrainer

Kupka

Increased translocator protein (TSPO) mRNA levels in colon but not in rectum carcinoma

European Surgical Research 2007 39

1076444

6 359 363

10.1159/000106380

2-s2.0-35349011132

19.

Maaser

Grabowski

Sutter

A. P.

Höpfner

Foss

H. D.

Stein

Berger

Gavish

Zeitz

Scherübl

Overexpression of the peripheral benzodiazepine receptor is a relevant prognostic factor in stage III colorectal cancer

Clinical Cancer Research 2002 8

1076444

10 3205 3209

12374690

20.

Beinlich

Strohmeier

Kaufmann

Kuhl

Relation of cell proliferation to expression of peripheral benzodiazepine receptors in human breast cancer cell lines

Biochemical Pharmacology 2000 60

1076444

3 397 402

10.1016/S0006-2952(00)00325-7

2-s2.0-0034257080

10856435

21.

Galiegue

Casellas

Kramar

Tinel

Simony-Lafontaine

Immunohistochemical assessment of the peripheral benzodiazepine receptor in breast cancer and its relationship with survival

Clinical Cancer Research 2004 10

1076444

6 2058 2064

10.1158/1078-0432.CCR-03-0988

2-s2.0-1642334068

15041726

22.

Sutter

A. P.

Maaser

Höpfner

Barthel

Grabowski

Faiss

Carayon

Zeitz

Scherübl

Specific ligands of the peripheral benzodiazepine receptor induce apoptosis and cell cycle arrest in human esophageal cancer cells

International Journal of Cancer 2002 102

1076444

4 318 327

10.1002/ijc.10724

2-s2.0-0036888292

12402299

23.

Batra

Iosif

C. S.

Peripheral benzodiazepine receptor in human endometrium and endometrial carcinoma

Anticancer Research 2000 20

1076444

1A 463 466

10769697

24.

Venturini

Alho

Podkletnova

Corsi

Rybnikova

Pellicci

Baraldi

Pelto-Huikko

Helén

Zeneroli

M. L.

Increased expression of peripheral benzodiazepine receptors and diazepam binding inhibitor in human tumors sited in the liver

Life Sciences 1999 65

1076444

21 2223 2231

10.1016/S0024-3205(99)00487-7

2-s2.0-0033569593

10576594

25.

Grande

Ljungman

P. L. S.

Eneroth

Bellander

Rizzuto

Association between cardiovascular disease and long-term exposure to air pollution with the risk of dementia

JAMA Neurology 2020 77

1076444

7 801 809

10.1001/jamaneurol.2019.4914

32227140

26.

Power

M. C.

Weisskopf

M. G.

Alexeeff

S. E.

Coull

B. A.

Spiro

III. Schwartz

Traffic-related air pollution and cognitive function in a cohort of older men

Environmental Health Perspectives 2011 119

1076444

5 682 687

10.1289/ehp.1002767

2-s2.0-79955811851

21172758

27.

Ritz

Lee

P. C.

Hansen

Lassen

C. F.

Ketzel

Sørensen

Raaschou-Nielsen

Traffic-related air pollution and Parkinson's disease in Denmark: a case-control study

Environmental Health Perspectives 2016 124

1076444

3 351 356

10.1289/ehp.1409313

2-s2.0-84959264593

26151951

28.

Wang

Eliot

M. N.

Wellenius

G. A.

Short-term changes in ambient particulate matter and risk of stroke: a systematic review and meta-analysis

Journal of the American Heart Association 2014 3

1076444

10.1161/JAHA.114.000983

2-s2.0-84926351906

25103204

29.

Kim

K. N.

Lim

Y. H.

Bae

H. J.

Kim

Jung

Hong

Y. C.

Long-term fine particulate matter exposure and major depressive disorder in a community-based urban cohort

Environmental Health Perspectives 2016 124

1076444

10 1547 1553

10.1289/EHP192

2-s2.0-84989332093

27129131

30.

Kioumourtzoglou

M. A.

Power

M. C.

Hart

J. E.

Okereke

O. I.

Coull

B. A.

Laden

Weisskopf

M. G.

The association between air pollution and onset of depression among middle-aged and older women

American Journal of Epidemiology 2017 185

1076444

9 801 809

10.1093/aje/kww163

2-s2.0-85020235204

28369173

31.

Block

M. L.

Zecca

Hong

J. S.

Microglia-mediated neurotoxicity: uncovering the molecular mechanisms

Nature Reviews. Neuroscience 2007 8

1076444

1 57 69

10.1038/nrn2038

2-s2.0-33845768784

17180163

32.

Campbell

Oldham

Becaria

Bondy

S. C.

Meacher

Sioutas

Misra

Mendez

L. B.

Kleinman

Particulate matter in polluted air may increase biomarkers of inflammation in mouse brain

Neurotoxicology 2005 26

1076444

1 133 140

10.1016/j.neuro.2004.08.003

2-s2.0-7444238755

15527881

33.

Allen

J. L.

Liu

Weston

Prince

Oberdörster

Finkelstein

J. N.

Johnston

C. J.

Cory-Slechta

D. A.

Developmental exposure to concentrated ambient ultrafine particulate matter air pollution in mice results in persistent and sex-dependent behavioral neurotoxicity and glial activation

Toxicological Sciences 2014 140

1076444

1 160 178

10.1093/toxsci/kfu059

2-s2.0-84903841144

24690596

34.

Block

M. L.

Calderon-Garciduenas

Air pollution: mechanisms of neuroinflammation and CNS disease

Trends in Neurosciences 2009 32

1076444

9 506 516

10.1016/j.tins.2009.05.009

2-s2.0-69949111672

19716187

35.

Chen

M. K.

Guilarte

T. R.

Translocator protein 18 kDa (TSPO): molecular sensor of brain injury and repair

Pharmacology & Therapeutics 2008 118

1076444

1 1 17

10.1016/j.pharmthera.2007.12.004

2-s2.0-42649115741

18374421

36.

Choi

Ifuku

Noda

Guilarte

T. R.

Translocator protein (18 kDa)/peripheral benzodiazepine receptor specific ligands induce microglia functions consistent with an activated state

Glia 2011 59

1076444

2 219 230

10.1002/glia.21091

2-s2.0-78650234355

21125642

37.

Banati

R. B.

Goerres

G. W.

Myers

Gunn

R. N.

Turkheimer

F. E.

Kreutzberg

G. W.

Brooks

D. J.

Jones

Duncan

J. S.

[11C](R)-PK11195 positron emission tomography imaging of activated microglia in vivo in Rasmussen's encephalitis

Neurology 1999 53

1076444

9 2199 2203

10.1212/WNL.53.9.2199

10599809

38.

Chauveau

Boutin

Van Camp

Dolle

Tavitian

Nuclear imaging of neuroinflammation: a comprehensive review of [11C]PK11195 challengers

European Journal of Nuclear Medicine and Molecular Imaging 2008 35

1076444

12 2304 2319

10.1007/s00259-008-0908-9

2-s2.0-58149303036

18828015

39.

Pappata

Cornu

Samson

Prenant

Benavides

Scatton

Crouzel

Hauw

J. J.

Syrota

PET study of carbon-11-PK 11195 binding to peripheral type benzodiazepine sites in glioblastoma: a case report

Journal of Nuclear Medicine 1991 32

1076444

8 1608 1610

1651383

40.

Mizrahi

Rusjan

P. M.

Vitcu

Wilson

A. A.

Houle

Bloomfield

P. M.

Whole body biodistribution and radiation dosimetry in humans of a new PET ligand, [(18) F]-FEPPA, to image translocator protein (18 kDa)

Molecular Imaging and Biology 2013 15

1076444

3 353 359

10.1007/s11307-012-0589-4

2-s2.0-84877733306

22895910

41.

Rusjan

P. M.

Wilson

A. A.

Bloomfield

P. M.

Vitcu

Meyer

J. H.

Houle

Mizrahi

Quantitation of translocator protein binding in human brain with the novel radioligand [18F]-FEPPA and positron emission tomography

Journal of Cerebral Blood Flow and Metabolism 2011 31

1076444

8 1807 1816

10.1038/jcbfm.2011.55

2-s2.0-79960997910

21522163

42.

Vignal

Cisternino

Rizzo-Padoin

San

Hontonnou

Gelé

Declèves

Sarda-Mantel

Hosten

[(18)F]FEPPA a TSPO radioligand: optimized radiosynthesis and evaluation as a PET radiotracer for brain inflammation in a peripheral LPS-injected mouse model

Molecules 2018 23

1076444

6, article 1375

43.

Wilson

A. A.

Garcia

Parkes

McCormick

Stephenson

K. A.

Houle

Vasdev

Radiosynthesis and initial evaluation of [¹⁸F]-FEPPA for PET imaging of peripheral benzodiazepine receptors

Nuclear Medicine and Biology 2008 35

1076444

3 305 314

10.1016/j.nucmedbio.2007.12.009

2-s2.0-40749142849

18355686

44.

Fujimura

Ikoma

Yasuno

Suhara

Ota

Matsumoto

Nozaki

Takano

Kosaka

Zhang

M. R.

Nakao

Suzuki

Kato

Ito

Quantitative analyses of 18F-FEDAA1106 binding to peripheral benzodiazepine receptors in living human brain

Journal of Nuclear Medicine 2006 47

1076444

1 43 50

16391186

45.

Zhang

M. R.

Maeda

Furutsuka

Yoshida

Ogawa

Suhara

Suzuki

[¹⁸F]FMDAA1106 and [¹⁸F]FEDAA1106: two positron-emitter labeled ligands for peripheral benzodiazepine receptor (PBR)

Bioorganic & Medicinal Chemistry Letters 2003 13

1076444

2 201 204

10.1016/S0960-894X(02)00886-7

2-s2.0-0037212103

12482423

46.

Golla

S. S.

Boellaard

Oikonen

Hoffmann

van Berckel

B. N. M.

Windhorst

A. D.

Virta

Haaparanta-Solin

Luoto

Savisto

Solin

Valencia

Thiele

Eriksson

Schuit

R. C.

Lammertsma

A. A.

Rinne

J. O.

Quantification of [18F]DPA-714 binding in the human brain: initial studies in healthy controls and Alzheimer's disease patients

Journal of Cerebral Blood Flow and Metabolism 2015 35

1076444

5 766 772

10.1038/jcbfm.2014.261

2-s2.0-84928826319

25649991

47.

Peyronneau

M. A.

Saba

Goutal

Damont

Dollé

Kassiou

Bottlaender

Valette

Metabolism and quantification of [(18)F]DPA-714, a new TSPO positron emission tomography radioligand

Drug Metabolism and Disposition 2013 41

1076444

1 122 131

10.1124/dmd.112.046342

2-s2.0-84871602840

23065531

48.

Dickstein

L. P.

Zoghbi

S. S.

Fujimura

Imaizumi

Zhang

Pike

V. W.

Innis

R. B.

Fujita

Comparison of 18F- and 11C-labeled aryloxyanilide analogs to measure translocator protein in human brain using positron emission tomography

European Journal of Nuclear Medicine and Molecular Imaging 2011 38

1076444

2 352 357

10.1007/s00259-010-1622-y

2-s2.0-79551569584

21085954

49.

Fujimura

Zoghbi

S. S.

Simeon

F. G.

Taku

Pike

V. W.

Innis

R. B.

Fujita

Quantification of translocator protein (18 kDa) in the human brain with PET and a novel radioligand, (18)F-PBR06

Journal of Nuclear Medicine 2009 50

1076444

7 1047 1053

10.2967/jnumed.108.060186

2-s2.0-67650088553

19525468

50.

Ory

Celen

Verbruggen

Bormans

PET radioligands for in vivo visualization of neuroinflammation

Current Pharmaceutical Design 2014 20

1076444

37 5897 5913

10.2174/1381612820666140613120212

2-s2.0-84911485957

24939192

51.

Zhang

Shao

Hou

Zhang

Josephson

Meyer

J. H.

Zhang

M. R.

Vasdev

Wang

Liang

S. H.

Recent developments on PET radiotracers for TSPO and their applications in neuroimaging

Acta Pharmaceutica Sinica B 2021 11

1076444

2 373 393

10.1016/j.apsb.2020.08.006

33643818

52.

Chau

W. F.

Black

A. M.

Clarke

Durrant

Gausemel

Khan

Mantzilas

Oulie

Rogstad

Trigg

Jones

P. A.

Exploration of the impact of stereochemistry on the identification of the novel translocator protein PET imaging agent [¹⁸F]GE-180

Nuclear Medicine and Biology 2015 42

1076444

9 711 719

10.1016/j.nucmedbio.2015.05.004

2-s2.0-84937728479

26072270

53.

Fan

Calsolaro

Atkinson

R. A.

Femminella

G. D.

Waldman

Buckley

Trigg

Brooks

D. J.

Hinz

Edison

Flutriciclamide (18F-GE180) PET: first-in-human PET study of novel third-generation in vivo marker of human translocator protein

Journal of Nuclear Medicine 2016 57

1076444

11 1753 1759

10.2967/jnumed.115.169078

2-s2.0-84994819090

27261523

54.

Wickstrøm

Clarke

Gausemel

Horn

Jørgensen

Khan

Mantzilas

Rajanayagam

In't Veld

D. J.

Trigg

The development of an automated and GMP compliant FASTlab synthesis of [(18) F]GE-180; a radiotracer for imaging translocator protein (TSPO)

Journal of Labelled Compounds and Radiopharmaceuticals 2014 57

1076444

1 42 48

10.1002/jlcr.3112

2-s2.0-84892820634

24448744

55.

Langen

K. J.

Willuweit

TSPO PET using 18F-GE-180: a new perspective in neurooncology?

European Journal of Nuclear Medicine and Molecular Imaging 2017 44

1076444

13 2227 2229

10.1007/s00259-017-3838-6

2-s2.0-85031412262

29026945

56.

Zanotti-Fregonara

Veronese

Pascual

Rostomily

R. C.

Turkheimer

Masdeu

J. C.

The validity of 18F-GE180 as a TSPO imaging agent

European Journal of Nuclear Medicine and Molecular Imaging 2019 46

1076444

6 1205 1207

10.1007/s00259-019-4268-4

2-s2.0-85060256421

30656358

57.

Zanotti-Fregonara

Pascual

Rizzo

Pal

Beers

Carter

Appel

S. H.

Atassi

Masdeu

J. C.

Head-to-head comparison of11C-PBR28 and18F-GE180 for quantification of the translocator protein in the human brain

Journal of Nuclear Medicine 2018 59

1076444

8 1260 1266

10.2967/jnumed.117.203109

2-s2.0-85042671070

29348317

58.

Vasdev

Green

D. E.

Vines

D. C.

McLarty

McCormick

P. N.

Moran

M. D.

Houle

Wilson

A. A.

Reilly

R. M.

Positron-emission tomography imaging of the TSPO with [(18)F]FEPPA in a preclinical breast cancer model

Cancer Biotherapy & Radiopharmaceuticals 2013 28

1076444

3 254 259

10.1089/cbr.2012.1196

2-s2.0-84876143204

23350894

59.

Zammit

Tao

Olsen

M. E.

Metzger

Vermilyea

S. C.

Bjornson

Slesarev

Block

W. F.

Fuchs

Phillips

Bondarenko

Zhang

S. C.

Emborg

M. E.

Christian

B. T.

[18F]FEPPA PET imaging for monitoring CD68-positive microglia/macrophage neuroinflammation in nonhuman primates

EJNMMI Research 2020 10

1076444

1 1 12

10.1186/s13550-020-00683-5

60.

Attwells

Setiawan

Rusjan

P. M.

Hutton

Rafiei

Varughese

Kahn

Kish

S. J.

Vasdev

Houle

Translocator protein distribution volume predicts reduction of symptoms during open-label trial of celecoxib in major depressive disorder

Biological Psychiatry 2020 88

1076444

8 649 656

10.1016/j.biopsych.2020.03.007

61.

Attwells

Setiawan

Wilson

A. A.

Rusjan

P. M.

Mizrahi

Miler

Richter

M. A.

Kahn

Kish

S. J.

Houle

Ravindran

Meyer

J. H.

Inflammation in the neurocircuitry of obsessive-compulsive disorder

JAMA Psychiatry 2017 74

1076444

8 833 840

10.1001/jamapsychiatry.2017.1567

2-s2.0-85027483326

28636705

62.

Da Silva

Hafizi

Watts

J. J.

Weickert

C. S.

Meyer

J. H.

Houle

Rusjan

Mizrahi

In vivo imaging of translocator protein in long-term Cannabis users

JAMA Psychiatry 2019 76

1076444

12 1305 1313

10.1001/jamapsychiatry.2019.2516

2-s2.0-85072397295

31532458

63.

Hafizi

Tseng

H. H.

Rao

Selvanathan

Kenk

Bazinet

R. P.

Suridjan

Wilson

A. A.

Meyer

J. H.

Remington

Houle

Rusjan

P. M.

Mizrahi

Imaging microglial activation in untreated first-episode psychosis: a PET study with [18F]FEPPA

The American Journal of Psychiatry 2017 174

1076444

2 118 124

10.1176/appi.ajp.2016.16020171

2-s2.0-85011559339

27609240

64.

Hartimath

S. V.

Khanapur

Boominathan

Jiang

Cheng

Yong

F. F.

Tan

P. W.

Robins

E. G.

Goggi

J. L.

Imaging adipose tissue browning using the TSPO-18kDa tracer [¹⁸F]FEPPA

Molecular Metabolism 2019 25

1076444

154 158

10.1016/j.molmet.2019.05.003

2-s2.0-85065622751

31105057

65.

Setiawan

Wilson

A. A.

Mizrahi

Rusjan

P. M.

Miler

Rajkowska

Suridjan

Kennedy

J. L.

Rekkas

P. V.

Houle

Meyer

J. H.

Role of translocator protein density, a marker of neuroinflammation, in the brain during major depressive episodes

JAMA Psychiatry 2015 72

1076444

3 268 275

10.1001/jamapsychiatry.2014.2427

2-s2.0-84925357658

25629589

66.

Yilmaz

Strafella

A. P.

Bernard

Schulte

van den Heuvel

Schneiderhan-Marra

Knorpp

Joos

T. O.

Leypoldt

Geritz

Hansen

Heinzel

Apel

Gasser

Lang

A. E.

Berg

Maetzler

Marras

Serum inflammatory profile for the discrimination of clinical subtypes in Parkinson's disease

Frontiers in Neurology 2018 9

1076444

1123

10.3389/fneur.2018.01123

30622507

67.

USP

Monographs: fludeoxyglucose F 18 injection

2015

3964

68.

Lee

S. H.

Lee

P. H.

Liang

H. J.

Tang

C. H.

Chen

T. F.

Cheng

T. J.

Lin

C. Y.

Brain lipid profiles in the spontaneously hypertensive rat after subchronic real-world exposure to ambient fine particulate matter

Science of The Total Environment 2020 707

1076444

135603

10.1016/j.scitotenv.2019.135603

31784156

69.

Yan

Y. H.

Chou

C. C. K.

Wang

J. S.

Tung

C. L.

Y. R.

Cheng

T. J.

Subchronic effects of inhaled ambient particulate matter on glucose homeostasis and target organ damage in a type 1 diabetic rat model

Toxicology and Applied Pharmacology 2014 281

1076444

2 211 220

10.1016/j.taap.2014.10.005

2-s2.0-84912092069

70.

Chuang

H. C.

Chen

H. C.

Chai

P. J.

Liao

H. T.

C. F.

Chen

C. L.

Jhan

M. K.

Hsieh

H. I.

K. Y.

Chen

T. F.

Cheng

T. J.

Neuropathology changed by 3- and 6-months low-level PM2.5 inhalation exposure in spontaneously hypertensive rats

Particle and Fibre Toxicology 2020 17

1076444

1 12

71.

Boellaard

Standards for PET image acquisition and quantitative data analysis

Journal of Nuclear Medicine 2009 50

1076444

Suppl 1 11S 20S

10.2967/jnumed.108.057182

2-s2.0-66149106325

19380405

72.

Bai

K. J.

Chuang

K. J.

Chen

C. L.

Jhan

M. K.

Hsiao

T. C.

Cheng

T. J.

Chang

L. T.

Chang

T. Y.

Chuang

H. C.

Microglial activation and inflammation caused by traffic-related particulate matter

Chemico-Biological Interactions 2019 311

1076444

108762

10.1016/j.cbi.2019.108762

2-s2.0-85069815216

31348917

73.

Berroterán-Infante

Balber

Furlinger

Bergmann

Lanzenberger

Hacker

Mitterhauser

Wadsak

[18F]FEPPA: improved automated radiosynthesis, binding affinity, and preliminary in vitro evaluation in colorectal cancer

ACS Medicinal Chemistry Letters 2018 9

1076444

3 177 181

10.1021/acsmedchemlett.7b00367

2-s2.0-85043481813

29541356

74.

Chang

C. W.

Chiu

C. H.

Lin

M. H.

H. M.

T. H.

Wang

P. Y.

Kuo

Y. Y.

Huang

Y. Y.

Shiue

C. Y.

Huang

W. S.

Yeh

S. H. H.

GMP-compliant fully automated radiosynthesis of [18F]FEPPA for PET/MRI imaging of regional brain TSPO expression

EJNMMI Research 2021 11

1076444

1 26

10.1186/s13550-021-00768-9

33725191

75.

Narayanaswami

Tong

Schifani

Bloomfield

P. M.

Dahl

Vasdev

Preclinical evaluation of TSPO and MAO-B PET radiotracers in an LPS model of neuroinflammation

PET Clinics 2021 16

1076444

2 233 247

10.1016/j.cpet.2020.12.003

33648665

76.

Young

Y. W.

Barback

C. V.

Stolz

L. A.

Groman

S. M.

Vera

D. R.

Hoh

Kotta

K. K.

Minassian

Powell

S. B.

Brody

A. L.

MicroPET evidence for a hypersensitive neuroinflammatory profile of gp120 mouse model of HIV

Psychiatry Research: Neuroimaging 2022 321

1076444

111445

10.1016/j.pscychresns.2022.111445

35101828

77.

Setiawan

Attwells

Wilson

A. A.

Mizrahi

Rusjan

P. M.

Miler

Sharma

Kish

Houle

Meyer

J. H.

Association of translocator protein total distribution volume with duration of untreated major depressive disorder: a cross-sectional study

Lancet Psychiatry 2018 5

1076444

4 339 347

10.1016/S2215-0366(18)30048-8

2-s2.0-85042592268

29496589

78.

Kenk

Selvanathan

Rao

Suridjan

Rusjan

Remington

Meyer

J. H.

Wilson

A. A.

Houle

Mizrahi

Imaging neuroinflammation in gray and white matter in schizophrenia: an in-vivo PET study with [18F]-FEPPA

Schizophrenia Bulletin 2015 41

1076444

1 85 93

10.1093/schbul/sbu157

2-s2.0-84928798486

25385788

79.

Suridjan

Pollock

B. G.

Verhoeff

N. P.

Voineskos

A. N.

Chow

Rusjan

P. M.

Lobaugh

N. J.

Houle

Mulsant

B. H.

Mizrahi

_In-vivo_ imaging of grey and white matter neuroinflammation in Alzheimer 's disease: a positron emission tomography study with a novel radioligand, [¹⁸F]-FEPPA

Molecular Psychiatry 2015 20

1076444

12 1579 1587

10.1038/mp.2015.1

2-s2.0-84947615776

25707397

80.

Al-Khishman

N. U.

Roseborough

A. D.

Levit

Allman

B. L.

Anazodo

U. C.

Fox

M. S.

Whitehead

S. N.

Thiessen

J. D.

TSPO PET detects acute neuroinflammation but not diffuse chronically activated MHCII microglia in the rat

EJNMMI Research 2020 10

1076444

1 1 10

10.1186/s13550-020-00699-x

32990808

Neuroinflammation in Low-Level PM2.5-Exposed Rats Illustrated by PET via an Improved Automated Produced [ 18 F]FEPPA: A Feasibility Study

Abstract

1. Introduction

2. Materials and Methods

2.1. Automated Radiosynthesis

2.2. Quality Control (QC) and Stability Tests of the [18F]FEPPA (2)

2.3. MicroPET Imaging of the [18F]FEPPA (2) Injection in Rats Exposed to Ambient Fine Particulate Matter (PM2.5)

2.3.1. Animals

2.4. Immunohistochemical Analysis

2.5. Statistical Analysis

3. Results

3.1. A One-Pot Automated Synthesis of the [18F]FEPPA (2) Using a Modified FxFN Module

3.2. MicroPET Imaging of the [18F]FEPPA (2) Injection in Rats Exposed to Ambient Fine Particulate Matter (PM2.5)

3.3. Immunohistochemical (IHC) Staining of the Rat Brain

4. Discussion

5. Conclusions

Footnotes

Data Availability

Ethical Approval

Conflicts of Interest

Authors’ Contributions

Acknowledgments

Supplementary Materials

References

2.2. Quality Control (QC) and Stability Tests of the [¹⁸F]FEPPA (2)

2.3. MicroPET Imaging of the [¹⁸F]FEPPA (2) Injection in Rats Exposed to Ambient Fine Particulate Matter (PM2.5)

3.1. A One-Pot Automated Synthesis of the [¹⁸F]FEPPA (2) Using a Modified Fx_FN Module

3.2. MicroPET Imaging of the [¹⁸F]FEPPA (2) Injection in Rats Exposed to Ambient Fine Particulate Matter (PM2.5)