spr 322c  (StressMarq)

Journal: Neural Regeneration Research

Article Title: Low-density lipoprotein receptor–related protein 1 mediates α-synuclein transmission from the striatum to the substantia nigra in animal models of Parkinson’s disease

doi: 10.4103/NRR.NRR-D-23-01965

Figure Lengend Snippet: LRP1 expression is increased in the nigrostriatal system of a monkey model of PD induced by α-syn PFFs injection. (A) Clinical rating score of monkeys after stereotactic injection of 600 μg of α-syn PFFs or an equal volume of normal saline for 4 months. (B) Immunohistochemistry detection of TH in the STR (left) and SN (right). The TH signal intensity in the STR and the number of TH-positive neurons in the SN were lower in the PFF group. The black arrow heads indicate typical TH-positive neurons. Scale bars: 100 μm. (C) Quantitative immunohistochemistry density analysis of TH in the STR. (D) Quantitation of the ratio of TH-positive neurons in the SN in the PFF group compared with the Sham group. (E) Western blot analysis of TH, LRP1, and α-syn expression levels in the monkey STR. (F–H) Densitometric analysis of TH (F), LRP1 (G), and α-syn (H) expression levels in the STR. (I) Western blot analysis of TH, LRP1, and α-syn expression levels in the monkey SN. (J–L) Densitometric analysis of TH (J), LRP1 (K), and α-syn (L) expression levels in the SN. Data are expressed as mean ± SD ( n = 3). * P < 0.05, ** P < 0.01, *** P < 0.001, vs. Sham group (unpaired t -test). LRP1: Low-density lipoprotein receptor-related protein 1; PD: Parkinson’s disease; PFF: pre-formed fibril; SN: substantia nigra; STR: striatum; TH: tyrosine hydroxylase; α-syn: α-synuclein.

Article Snippet: α-Syn PFFs (SPR-322, StressMarq, Victoria, Canada) were sonicated with a handheld probe (VCX130, SONICS & MATERIALS, Newtown, CT, USA) as follows: amplitude 30% and time 15 seconds (pulse on 3 seconds, pulse off 6 seconds) (Volpicelli-Daley et al., 2014; Sun et al., 2022b).

Techniques: Expressing, Injection, Saline, Immunohistochemistry, Quantitation Assay, Western Blot

Journal: Neural Regeneration Research

Article Title: Low-density lipoprotein receptor–related protein 1 mediates α-synuclein transmission from the striatum to the substantia nigra in animal models of Parkinson’s disease

doi: 10.4103/NRR.NRR-D-23-01965

Figure Lengend Snippet: LRP1 expression is increased in the nigrostriatal system of a mouse model of PD induced by injection with α-Syn PFFs. (A) Visual gait analysis of walking, gait, and footprint pressure in mice after stereotactic injection with 5 μg of α-Syn PFFs or an equal volume of PBS for 4 weeks ( n = 6). (B) Normal step sequence in mice ( n = 6). (C) Average speed in mice ( n = 6). (D) Tripod support time in mice ( n = 6). (E) Immunohistochemistry staining for TH in the STR. The images in the second row are enlarged images of the areas indicated in black boxes in the upper row. The TH signal intensity in the STR was lower in the PFF group. Scale bars: 200 μm, 40 μm (enlarged). (F) Immunohistochemistry staining for TH in the SN. The images in the second row are enlarged images of the areas indicated in black boxes in the upper row, and the white arrowheads indicate typical TH-positive neurons. There were fewer TH-positive neurons in the SN in the PFF group. Scale bars: 100 μm, 20 μm (enlarged). (G) Quantitative immunohistochemistry density analysis of TH in the STR, compared with the Sham group ( n = 3). (H) Quantitation of the ratio of TH-positive neurons in the SN in the PFF group compared with the Sham group ( n = 3). (I) Western blot analysis of TH, LRP1, and α-Syn expression levels in the mouse STR after injection with 5 μg of α-Syn PFFs or an equal volume of PBS ( n = 3). (J–L) Densitometric analysis of TH (J), LRP1 (K), and α-Syn (L) expression levels in the STR. (M) Western blot analysis of TH, LRP1, and α-Syn expression levels in the mouse SN ( n = 3). (N–P) Densitometric analysis of TH (N), LRP1 (O), and α-Syn (P) expression levels in the SN ( n = 3). Data are expressed as mean ± SD ( n = 3). * P < 0.05, ** P < 0.01, *** P < 0.001, vs . Sham group (unpaired t -test). LRP1: Low-density lipoprotein receptor-related protein 1; PBS: phosphate buffered solution; PD: Parkinson’s disease; PFF: pre-formed fibril; SN: substantia nigra; STR: striatum; TH: tyrosine hydroxylase; α-Syn: α-synuclein.

Article Snippet: α-Syn PFFs (SPR-322, StressMarq, Victoria, Canada) were sonicated with a handheld probe (VCX130, SONICS & MATERIALS, Newtown, CT, USA) as follows: amplitude 30% and time 15 seconds (pulse on 3 seconds, pulse off 6 seconds) (Volpicelli-Daley et al., 2014; Sun et al., 2022b).

Techniques: Expressing, Injection, Sequencing, Immunohistochemistry, Staining, Quantitation Assay, Western Blot

Journal: Neural Regeneration Research

Article Title: Low-density lipoprotein receptor–related protein 1 mediates α-synuclein transmission from the striatum to the substantia nigra in animal models of Parkinson’s disease

doi: 10.4103/NRR.NRR-D-23-01965

Figure Lengend Snippet: Exogenous α-syn PFFs upregulate LRP1 expression in PC12 cells. (A) Viability of PC12 cells incubated with different doses (0, 5, 10, 20, or 50 µg/mL) of α-Syn PFFs for 24 hours. ** P < 0.01, *** P < 0.001, vs . 0 µg/mL group. (B) Western blot analysis of LRP1 and α-syn expression levels in PC12 cells. (C, D) Densitometric analysis of LRP1 (C) and α-syn (D) expression levels. (E) Immunofluorescence analysis of LRP1 (green, Alexa Fluor 488), α-syn (red, Alexa Fluor 594), and DAPI (blue) expression. The LRP1 and α-syn signal intensities were stronger in PFF group than in Con group. Scale bars: 40 μm. (F, G) Quantitative immunofluorescence intensity analysis of LRP1 (F) and α-syn (G). * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001, vs . Con group; # P < 0.05, ## P < 0.01, vs. monomer group. Data are expressed as mean ± SD ( n = 3). Con: Control; DAPI: 4′,6-diamidino-2-phenylindole; PFF: pre-formed fibril; LRP1: low-density lipoprotein receptor-related protein 1; α-syn: α-synuclein.

Article Snippet: α-Syn PFFs (SPR-322, StressMarq, Victoria, Canada) were sonicated with a handheld probe (VCX130, SONICS & MATERIALS, Newtown, CT, USA) as follows: amplitude 30% and time 15 seconds (pulse on 3 seconds, pulse off 6 seconds) (Volpicelli-Daley et al., 2014; Sun et al., 2022b).

Techniques: Expressing, Incubation, Western Blot, Immunofluorescence, Control

Journal: Neural Regeneration Research

Article Title: Low-density lipoprotein receptor–related protein 1 mediates α-synuclein transmission from the striatum to the substantia nigra in animal models of Parkinson’s disease

doi: 10.4103/NRR.NRR-D-23-01965

Figure Lengend Snippet: LRP1 knockdown rescues the dopaminergic damage induced by exogenous α-syn PFFs. (A, B) Western blot analysis (A) and densitometric analysis (B) of TH expression levels in the STR of mice treated with PFFs. (C, D) Western blot analysis (C) and densitometric analysis (D) of TH expression levels in the SN of mice treated with PFFs. (E) Immunohistochemistry staining for TH in the STR of mice treated with PFFs and LRP1 siRNA. The decrease in relative TH intensity in STR observed in the PFF group was rescued by LRP1 siRNA treatment. The images in the second row are enlarged images of the areas indicated by black boxes in the upper row. Scale bars: 200 μm, 40 μm (enlarged). (F) Quantitative immunohistochemistry density analysis of TH in the STR. (G) Immunohistochemistry analysis of TH in the SN of mice treated with PFFs. The decreased ratio of TH-positive neurons in the SN of the Scramble + PFF group (PFF group) was rescued by LRP1 siRNA treatment. The images in the second row are enlarged images of the areas indicated by black boxes in the upper row, and the white arrowheads indicate typical TH-positive neurons. Scale bars: 100 μm, 20 μm (enlarged). (H) Quantitation of the ratio of TH-positive neurons in the SN compared with the PBS group. Data are expressed as mean ± SD ( n = 3). * P < 0.05, ** P < 0.01, *** P < 0.001, vs. Scramble + PBS group (PBS group); # P < 0.05, ## P < 0.01, vs. scramble + PFF group (PFF group). LRP1: Low-density lipoprotein receptor-related protein 1; PBS: phosphate buffered solution; PFF: pre-formed fibril; siRNA: small interfering RNA; SN: substantia nigra; STR: striatum; TH: tyrosine hydroxylase; α-syn: α-synuclein.

Article Snippet: α-Syn PFFs (SPR-322, StressMarq, Victoria, Canada) were sonicated with a handheld probe (VCX130, SONICS & MATERIALS, Newtown, CT, USA) as follows: amplitude 30% and time 15 seconds (pulse on 3 seconds, pulse off 6 seconds) (Volpicelli-Daley et al., 2014; Sun et al., 2022b).

Techniques: Knockdown, Western Blot, Expressing, Immunohistochemistry, Staining, Quantitation Assay, Small Interfering RNA

Journal: Neural Regeneration Research

Article Title: Low-density lipoprotein receptor–related protein 1 mediates α-synuclein transmission from the striatum to the substantia nigra in animal models of Parkinson’s disease

doi: 10.4103/NRR.NRR-D-23-01965

Figure Lengend Snippet: LRP1 suppression reduces the transmission of exogenous α-syn in vivo . (A) Immunofluorescence staining for LRP1 (green, Alexa Fluor 488), α-syn (red, Alexa Fluor 594), and DAPI (blue) in the mouse STR after stereotactic injection with α-syn PFFs for 6 weeks with or without LRP1 knockdown. The increase in the relative fluorescence intensity of LRP1 and α-syn in the STR of PFFs-treated mice was rescued by LRP1 siRNA treatment. Scale bars: 100 μm. (B, C) Quantitative immunofluorescence intensity of LRP1 (B) and α-syn (C) in mouse STR. (D) Immunofluorescence staining for LRP1 (green, Alexa Fluor 488), α-syn (red, Alexa Fluor 594), and DAPI (blue) in the SN of mice after stereotactic injection with α-syn PFFs for 6 weeks with or without LRP1 knockdown. The increase in the relative fluorescence intensity of LRP1 and α-syn in the SN of mice treated with PFFs was rescued by LRP1 siRNA treatment. The white arrowheads indicate typical cells exhibiting α-syn and LRP1 expression. Scale bars: 100 μm. (E, F) Quantitative immunofluorescence intensity of LRP1 (E) and α-syn (F) in the mouse SN. (G) Western blot analysis of LRP1 and α-syn expression levels in the mouse STR. (H, I) Densitometric analysis of LRP1 (H) and α-syn (I) expression levels in the mouse STR. (J) Western blot analysis of LRP1 and α-syn expression levels in the mouse SN. (K, L) Densitometric analysis of LRP1 (K) and α-syn (L) expression levels in the mouse SN. Data are expressed as mean ± SD ( n = 3). * P < 0.05, ** P < 0.01, vs. Scramble + PBS group (PBS group); # P < 0.05, ## P < 0.01, vs . Scramble + PFF group (PFF group). DAPI: 4′,6-Diamidino-2-phenylindole; LRP1: low-density lipoprotein receptor-related protein 1; PBS: phosphate buffered solution; PFF: pre-formed fibril; siRNA: small interfering RNA; SN: substantia nigra; STR: striatum; α-syn: α-synuclein.

Article Snippet: α-Syn PFFs (SPR-322, StressMarq, Victoria, Canada) were sonicated with a handheld probe (VCX130, SONICS & MATERIALS, Newtown, CT, USA) as follows: amplitude 30% and time 15 seconds (pulse on 3 seconds, pulse off 6 seconds) (Volpicelli-Daley et al., 2014; Sun et al., 2022b).

Techniques: Transmission Assay, In Vivo, Immunofluorescence, Staining, Injection, Knockdown, Fluorescence, Expressing, Western Blot, Small Interfering RNA

Journal: Neural Regeneration Research

Article Title: Low-density lipoprotein receptor–related protein 1 mediates α-synuclein transmission from the striatum to the substantia nigra in animal models of Parkinson’s disease

doi: 10.4103/NRR.NRR-D-23-01965

Figure Lengend Snippet: LRP1 mediates the uptake of α-syn PFFs by PC12 cells. (A) Western blot analysis of LRP1 and α-syn expression levels in PC12 cells with or without LRP1 knockdown (10 nM siRNA) after incubation with 10 µg/mL α-syn PFFs for 24 hours. (B, C) Densitometric analysis of LRP1 (B) and α-syn (C) expression. (D) Immunofluorescence staining for LRP1 (green, Alexa Fluor 488), α-syn (red, Alexa Fluor 594), and DAPI (blue). The increase in the relative fluorescence intensity levels of LRP1 and α-syn in the PFF group was rescued by LRP1 siRNA treatment. Scale bars: 40 μm. (E, F) Quantitative immunofluorescence intensity analysis of LRP1 (E) and α-syn (F). Data are expressed as mean ± SD ( n = 3). ** P < 0.01, *** P < 0.001, **** P < 0.0001, vs . Con group; # P < 0.05, ## P < 0.01, #### P < 0.0001, vs . PFF/– group. Con: Control; DAPI: 4′, 6-diamidino-2-phenylindole; PFF: pre-formed fibril; LRP1: low-density lipoprotein receptor-related protein 1; siRNA: small interfering RNA; α-syn: α-synuclein.

Article Snippet: α-Syn PFFs (SPR-322, StressMarq, Victoria, Canada) were sonicated with a handheld probe (VCX130, SONICS & MATERIALS, Newtown, CT, USA) as follows: amplitude 30% and time 15 seconds (pulse on 3 seconds, pulse off 6 seconds) (Volpicelli-Daley et al., 2014; Sun et al., 2022b).

Techniques: Western Blot, Expressing, Knockdown, Incubation, Immunofluorescence, Staining, Fluorescence, Control, Small Interfering RNA

Journal: Neural Regeneration Research

Article Title: Low-density lipoprotein receptor–related protein 1 mediates α-synuclein transmission from the striatum to the substantia nigra in animal models of Parkinson’s disease

doi: 10.4103/NRR.NRR-D-23-01965

Figure Lengend Snippet: Lysine residues in the α-syn N-terminus are vital for LRP1-mediated α-Syn internalization. (A) Heatmap of amino acids in different α-syn domains. (B) Western blot analysis of LRP1 and α-Syn levels in PC12 cells after addition of α-syn PFFs (10 µg/mL) with or without capping of lysine residues for 24 hours. (C, D) Densitometric analysis of LRP1 (C) and α-syn (D) expression levels. (E) Immunofluorescence staining for LRP1 (green, Alexa Fluor 488), α-Syn (red, Alexa Fluor 594), and DAPI (blue). The increase in the relative fluorescence intensity of LRP1 and α-syn in the PFF group was rescued by lysine capping of α-syn. Scale bars: 40 μm. (F, G) Quantitative immunofluorescence intensity analysis of LRP1 (F) and α-syn (G). Data are expressed as mean ± SD ( n = 3). * P < 0.05, ** P < 0.01, **** P < 0.0001, vs. Con group; # P < 0.05, ## P < 0.01, vs . PFF group. Con: Control; DAPI: 4′,6-diamidino-2-phenylindole; LRP1: low-density lipoprotein receptor-related protein 1; NHS: sulfo-NHS acetate; PFF: pre-formed fibril; α-syn: α-synuclein.

Article Snippet: α-Syn PFFs (SPR-322, StressMarq, Victoria, Canada) were sonicated with a handheld probe (VCX130, SONICS & MATERIALS, Newtown, CT, USA) as follows: amplitude 30% and time 15 seconds (pulse on 3 seconds, pulse off 6 seconds) (Volpicelli-Daley et al., 2014; Sun et al., 2022b).

Techniques: Western Blot, Expressing, Immunofluorescence, Staining, Fluorescence, Control

Journal: ACS Chemical Neuroscience

Article Title: Fluorescence Detection of Alpha-Synuclein Aggregates in the Gut Using a Peptide Probe

doi: 10.1021/acschemneuro.6c00069

Figure Lengend Snippet: Probe P1 shows high labeling accuracy for α-syn aggregates in vitro, compared to phosphorylated (pS129) α-syn antibody which was used as the gold standard reference staining for α-syn aggregates. (a) Structure of peptide-based probe P1 ; (b) Setup of primary hippocampal neuronal culture and induction of α-syn aggregation upon addition of PFFs; (c) Confocal microscopy of primary neuronal cells post 7 days of PFF addition. Nuclei are colored blue, total α-syn green, pS129 α-syn orange, and probe P1 in red. Scale bar represents 200 μm; (d) Probe P1 shows colocalized staining with the PS129 α-syn antibody staining. The probe labels most of the same focal locations, but with slightly larger surrounding area than the PS129 α-syn antibody hitsthis is still regarded as a positive colocalization. True positives (indicated by green arrows) are stained by both P1 and the pS129 α-syn antibody. False negatives (indicated by pink arrows) are not stained by P1 but stained by the pS129 α-syn antibody. Scale bar represents 100 μm; (e) Graph showing average positive predictive value (true positives/probe positives) of 87.4% and miss rate (false negatives/antibody positives) of 8.7%. Values on graph are given as the mean ± SEM calculated over 10 wells, with 9 fields of view each.

Article Snippet: Human recombinant α-syn monomers (SPR-321C), kinetically stable oligomers (SPR-484C), and PFFs (SPR-322C) were purchased from StressMarq Biosciences and used without further modification.

Techniques: Labeling, In Vitro, Staining, Confocal Microscopy

Journal: ACS Chemical Neuroscience

Article Title: Fluorescence Detection of Alpha-Synuclein Aggregates in the Gut Using a Peptide Probe

doi: 10.1021/acschemneuro.6c00069

Figure Lengend Snippet: Probe P1 exhibits preferential binding to α-syn fibrils over α-syn monomers. (a) Fluorescent excitation and (b) fluorescent emission spectra of P1 . 2 μM probe P1 was incubated with 10 μM of various α-syn species, including monomers, A* oligomers, B* oligomers, and PFFs. Concentrations of all α-syn species are expressed at their equivalent monomer concentration. Fluorescence intensity of P1 increases significantly upon interaction with PFFs (7.6× and 16× for excitation and emission respectively); (c) Fluorescence titration curve of P1 (0.5 μM) to PFFs, showing micromolar K D ; (d) Native-PAGE images of P1 with various α-syn species, showing that P1 preferentially binds to B* oligomers and PFFs, over α-syn monomers and A* oligomers. 2 μg of P1 was incubated with 2 μg of α-syn monomers and various α-syn aggregate species and run under native PAGE conditions. In-gel fluorescence was first visualized in the 647 channel, and then the gel was subsequently imaged in the brightfield channel after staining with Coomassie blue. Red bands correspond to P1 bound to the B* oligomers and PFFs; (e) Dot-blot images of P1 (0.5 μM) with various human α-syn species, showing concentration-dependent labeling of kinetically stable oligomers and PFFs.

Article Snippet: Human recombinant α-syn monomers (SPR-321C), kinetically stable oligomers (SPR-484C), and PFFs (SPR-322C) were purchased from StressMarq Biosciences and used without further modification.

Techniques: Binding Assay, Incubation, Concentration Assay, Fluorescence, Titration, Clear Native PAGE, Staining, Dot Blot, Labeling

Journal: Translational Psychiatry

Article Title: Impairment in stimulus-response learning as a cognitive biomarker in a model of synucleinopathy

doi: 10.1038/s41398-025-03795-5

Figure Lengend Snippet: A Schematic of α-synuclein pre-formed fibril production. α-synuclein type 1 pre-formed fibrils (PFF-1) were generated from monomers expressed in E.coli at room temperature overnight, while α-synuclein type 2 pre-formed fibrils (PFF-2) were generated from monomers expressed in E.coli grown at 37 °C for four hours. Following this, both fibril precursors underwent monomer purification and identical fibrilization protocols and were characterized in vitro. B SDS-PAGE of PFF-1 and PFF-2 after sedimentation revealed similar populations of insoluble (pellet) vs soluble (supernatant) materials, both consisting of predominantly insoluble species. C Transmission Electron Microscopy (TEM) of unsonicated PFF-1 and PFF-2 displayed similar architecture and range of fibril sizes, both appearing as elongated fibrils of mixed length. D Far UV circular dichroism (UV-CD) measurements identified a significant β-sheet presence in fibrils compared monomers with subtle differences between PFF-1 and PFF-2s. Thioflavin T (ThT) binding assay revealed a stronger binding capacity of PFF-1 (E) relative to PFF-2 (F) after fibrilization and during a seeding assay with additional monomer.

Article Snippet: Human aSyn PFF-1 and PFF-2 are available from a commercial source (StressMarq Biosciences Cat# SPR-322 and SPR-317, respectively), which are expressed, purified, and fibrilized using methods previously described [ , ].

Techniques: Generated, Purification, In Vitro, SDS Page, Sedimentation, Transmission Assay, Electron Microscopy, Circular Dichroism, Binding Assay

Journal: Translational Psychiatry

Article Title: Impairment in stimulus-response learning as a cognitive biomarker in a model of synucleinopathy

doi: 10.1038/s41398-025-03795-5

Figure Lengend Snippet: A M83 mice underwent stereotaxic surgery for inoculation of aSyn PFF-1, aSyn PFF-2 or PBS control into the right dorsal striatum. At 8 weeks post injection, they subsequently underwent a motor test battery, including assessments of motor strength, motor coordination and gait. Following completion, they were assessed for the cognitive ability to acquire stimulus-response associations, with the ‘Visuomotor Conditional Learning’ at 9-12 weeks post injection. At 16 weeks post-injection, they repeated the motor test battery, and then were processed for pathology and biochemistry. B Brain homogenates from aSyn PFF-1 and PFF-2 inoculated mice both displayed resistance to proteinase K but exhibited variable banding patterns upon digestion, as assessed by Western Blot using an antibody for total aSyn. C, D Furthermore, the same brain homogenates were evaluated by dotblots using an antibody with high affinity for aggregated aSyn (chBIIB054), revealing that while both fibril types exhibited significant binding, aSyn PFF-2 brain homogenates presented with a stronger signal (Welch’s ANOVA test, W 2, 5.34 = 7.03, p < 0.05, in which Welch-corrected unpaired t-tests revealed to be driven by a significant difference between PBS control and PFF-2 (p < 0.05). Graphics made with Biorender.com.

Article Snippet: Human aSyn PFF-1 and PFF-2 are available from a commercial source (StressMarq Biosciences Cat# SPR-322 and SPR-317, respectively), which are expressed, purified, and fibrilized using methods previously described [ , ].

Techniques: Control, Injection, Battery, Western Blot, Binding Assay

Journal: Translational Psychiatry

Article Title: Impairment in stimulus-response learning as a cognitive biomarker in a model of synucleinopathy

doi: 10.1038/s41398-025-03795-5

Figure Lengend Snippet: A aSyn PFF-1, PFF-2 or PBS control was unilaterally injected into the right dorsal striatum. Brains for immunohistochemistry (B, C) and immunoblots (D-I) were collected at 16 weeks post injection. B Immunostained brain slices imaged at 63X on a Leica Stellaris 5 Confocal Microscope for aSyn phosphorylated at S129 (green= pS129; blue= Hoescht), scale bar 25 µm. C Although inoculation was localized within the dorsal striatum, pathology spread to many areas including orbitofrontal cortex, piriform cortex, medulla, and substantia nigra. 40X, Leica Stellaris 5 Confocal Microscope, scale bar 50 µm. D–I To quantify this pathology further, protein extract from the striatum of PFF- and PBS-inoculated mice were extracted and quantified with immunoblots. D–F The levels of pS129 and total human aSyn found in the detergent-soluble (RIPA) fractions from PFF-1 and PFF-2 samples were expressed as the percentage fold change relative to PBS samples. No significant differences were found between PFF-1 and PFF-2 when comparing pS129 (unpaired t-test, p > 0.05) or human aSyn (unpaired t-test, p > 0.05). G–I The levels of pS129 and total human aSyn found in the detergent-insoluble fractions (UREA solubilization of RIPA-insoluble pellets) were expressed in the same way as the detergent-soluble fractions (RIPA). No significant differences were found between PFF-1 and PFF-2 when comparing pS129 (unpaired t-test, p > 0.05) or human aSyn (unpaired t-test, p > 0.05). Graphics made with Biorender.com.

Article Snippet: Human aSyn PFF-1 and PFF-2 are available from a commercial source (StressMarq Biosciences Cat# SPR-322 and SPR-317, respectively), which are expressed, purified, and fibrilized using methods previously described [ , ].

Techniques: Control, Injection, Immunohistochemistry, Western Blot, Microscopy

Journal: Translational Psychiatry

Article Title: Impairment in stimulus-response learning as a cognitive biomarker in a model of synucleinopathy

doi: 10.1038/s41398-025-03795-5

Figure Lengend Snippet: A Schematic of the forelimb grip force test, used to measure muscle strength. Subjects grasped a smooth, triangular pull bar with both forelimbs and the force exerted in Newtons (N) was measured. B aSyn PFF-2 but not PFF-1 inoculated mice displayed a significant reduction in forelimb muscle strength at 16 weeks post injection (WPI) relative to 8 (2-way RM ANOVA, Group x Session: main effect of group (F 2,72 = 4.409, p < 0.05), main effect of session (F 1,72 = 13.51, p < 0.001), and significant interaction (F 2,72 = 6.677, p < 0.01), in which Šídák’s multiple comparisons revealed to be driven by a significant difference within between 8 and 16 weeks within the PFF-2 group (adj. p < 0.0001). C Schematic of the accelerating Rota-rod, used to measure motor coordination and motor learning. Subjects were placed on a stationary rod which began to accelerate linearly from 5-36 revolutions per minute over 5 min, and latency to fall (s) was calculated automatically. D No significant difference between groups was found at 8 WPI (2-way RM ANOVA, Group x Session: main effect of session (F 9,675 = 15.41, p < 0.0001), but no main effect of group (p > 0.05). A significant interaction was first observed, (F 18, 675 = 1.767, p < 0.05), but Šídák’s multiple comparisons revealed no significant differences between groups (adj. p > 0.05)). However, (E) aSyn PFF-2 mice exhibited a shorter latency to fall at 16 WPI relative to aSyn PFF-1 and PBS controls (2-way RM ANOVA, Group x Session: Main effect of group (F 2,71 = 7.436, p < 0.01), main effect of session (F 9,639 = 3.205, p < 0.001), but no interaction (p > 0.05). To examine this main effect further, Šídák’s multiple comparisons revealed a significant difference between aSyn PFF-2 and PBS controls (adj. p < 0.01) and aSyn PFF-2 and PFF-1 (adj. p < 0.01)). F Schematic of the catwalk XT Gait Analysis, used to measure gait and locomotion. Footprints were captured while subjects voluntarily traversed a glass-plated CatWalk runway, towards a dark goal box at the end. G A significant difference between 8 and 16 weeks was revealed in the stride length of aSyn PFF-2 inoculated mice (2-way RM ANOVA, Group x Session: no main effect of group (p > 0.05), but a main effect of session (F 1,24 = 4.493, p < 0.05), and significant interaction (F 2,24= 3.646 , p < 0.05), in which Šídák’s multiple comparisons revealed a significant difference between timepoints within the aSyn PFF-2 group (adj. p < 0.01). H A significant difference between 8 and 16 weeks was revealed in the swing speed of aSyn PFF-2 inoculated mice (2-way RM ANOVA, Group x Session: no main effect of group (p > 0.05), but a main effect of session (F 1,24= 6.537 , p < 0.05), and significant interaction (F 2,24= 4.300 , p < 0.05) in which Šídák’s multiple comparisons revealed a significant difference between timepoints within the aSyn PFF-2 group (adj. p < 0.01). I A significant difference between 8 and 16 weeks was revealed in the step cycle of aSyn PFF-2 inoculated mice (2-way RM ANOVA, Group x Session: no main effect of group (p > 0.05), but a main effect of session (F 1,24= 9.232 , p < 0.01) and significant interaction (F 2,24= 4.035 , p < 0.05) in which Šídák’s multiple comparisons revealed a significant difference between timepoints within the aSyn PFF-2 group (adj. p < 0.01). Data presented as Mean + SEM, group x session Two-way RM ANOVA, ** p < 0.01, **** p < 0.0001. Graphics made with Biorender.com.

Article Snippet: Human aSyn PFF-1 and PFF-2 are available from a commercial source (StressMarq Biosciences Cat# SPR-322 and SPR-317, respectively), which are expressed, purified, and fibrilized using methods previously described [ , ].

Techniques: Injection

Journal: Translational Psychiatry

Article Title: Impairment in stimulus-response learning as a cognitive biomarker in a model of synucleinopathy

doi: 10.1038/s41398-025-03795-5

Figure Lengend Snippet: A Schematic of Visuomotor Conditional Learning (VMCL) task. Cognitive assessments were conducted within touchscreen systems equipped with a touch-sensitive screen, a reward magazine attached to a reward pump for delivery of strawberry milkshake liquid reward and ABET cognition software ( above ). VMCL was employed to evaluate the acquisition of stimulus-response (S-R) contingencies, with subjects learning the conditional rule: “if visual stimulus A is presented, make motor response to the right-flanking window; if visual stimulus B is presented, make motor response to the flight-flanking window” ( below ). All subjects underwent VMCL testing for 20 sessions, 5-7 sessions per week. B M83 mice inoculated with aSyn PFF-1 and PFF-2 were significantly impaired at acquiring VMCL, as demonstrated by lower percent correct responses relative to PBS-inoculated mice (2-way RM ANOVA, Group x Session: main effect of group (F 2,60=10.53 , p < 0.001), main effect of session (F 2.686, 161.2=10.53 , p < 0.0001) and significant interaction (F 8,240=3.131 , p < 0.01), in which Šídák’s multiple comparisons revealed to be driven by a significant difference between PBS control and PFF-1 in sessions 2 (adj. p < 0.05), 4 (adj. p < 0.05) and 5 (adj. p < 0.05), and a significant difference between PBS control and PFF-2 in sessions 2 (adj. p < 0.01), 3 (adj. p < 0.001), 4 (adj. p < 0.01), and 5 (adj. p < 0.01)). C While all groups began at chance in Block 1 (1-way ANOVA, p > 0.05), (D) both aSyn-inoculated groups were significantly impaired relative to controls in Block 5 (1-way ANOVA, F 8.764 , p < 0.001, in which Šídák’s multiple comparisons revealed to be driven by a significant difference between PBS controls and PFF-1 (adj. p < 0.05), and between PBS controls PFF-2 (adj. p < 0.001)). E No significant difference was found in the percentage of missed trials across groups (RM Mixed-Effects Model, Group x Session: main effect of session (F 2.050,118.4= 20.55 , p < 0.0001), but no main effect of group or interaction (p > 0.05)), (F) but aSyn PFF-1 and PFF-2 mice exhibited a greater number of correction trials (2-way RM ANOVA, Group x Session: main effect of group (F 2,60= 6.008 , p < 0.01), main effect of session (F 3.240,194.4= 66.22 , p < 0.0001) but no interaction (p > 0.05)), and (G) an elevated perseveration index compared to controls (RM Mixed-Effects Model, Group x Session: main effect of group (F 2,60=4.648 , p < 0.05), main effect of session (F 3.191,171.5=24.48 , p < 0.0001), but no interaction (p > 0.05)). Furthermore, comparing task latencies, no significant difference was found in the latency to make correct choices (H) (2-way RM ANOVA: Group x Session: p > 0.05), but aSyn PFF-1 and PFF-2 mice took significantly longer to make incorrect choices (I) across VMCL acquisition compared to PBS controls (2-way RM ANOVA: Group x Session: main effect of group (F 2,60=5.384 , p < 0.01), main effect of session (F 3.202,192.1= 62.37 , p < 0.0001), and significant interaction (F 8,240=1.985 , p < 0.05), in which Šídák’s multiple comparisons revealed to be driven by a significant difference between PBS control and PFF-1 in sessions 2 (adj. p < 0.01), 3 (adj. p < 0.01) and 5 (adj. p < 0.05), and a significant difference between PBS control and PFF-2 in sessions 3 (adj. p < 0.05) and 5 (adj. p < 0.05)). No significant difference was found for the latency to collect rewards (J) (2-way RM ANOVA, Group x Session: main effect of session (F 2.275,135.9=17.67 , p < 0.0001), but no main effect of group or interaction (p > 0.05)). Data presented as Mean + SEM, * p < 0.05, ** p < 0.01, *** p < 0.001. Graphics made with Biorender.com.

Article Snippet: Human aSyn PFF-1 and PFF-2 are available from a commercial source (StressMarq Biosciences Cat# SPR-322 and SPR-317, respectively), which are expressed, purified, and fibrilized using methods previously described [ , ].

Techniques: Software, Control, Blocking Assay