Increased p-cresol levels reportedly alter brain dopamine metabolism and exacerbate neurological disorders in experimental animals. In contrast to toxic concentrations, low doses of p-cresol may have distinct effects on neuronal metabolism. However, the role of p-cresol in synapse remodeling, neurite outgrowth, and other anabolic processes in neurons remains elusive. We propose that low doses of p-cresol affect neuronal cell structural remodeling compared with the high concentration-mediated harmful effects. Thus, the effects of p-cresol on the secretion of brain-derived neurotrophic factor (BDNF) and neurofilament subunit expression were examined using rat pheochromocytoma cells (PC-12 cells). We observed that low doses of p-cresol potentiated nerve growth factor-induced differentiation via secretion of BDNF in cultured PC-12 cells. Opioidergic compounds modulated these p-cresol effects, which were reversed by oxytocin. We propose that this effect of p-cresol has an adaptive and compensatory character and can be attributed to the induction of oxidative stress. Accordingly, we hypothesize that low doses of p-cresol induce mild oxidative stress, stimulating BDNF release by activating redox-sensitive genes. Given that the intestinal microbiome is the primary source of endogenous p-cresol, the balance between gut microbiome strains (especially Clostridium species) and opioidergic compounds may directly influence neuroplasticity.
Citation: Gigi Tevzadze, Tamar Barbakadze, Elisabed Kvergelidze, Elene Zhuravliova, Lali Shanshiashvili, David Mikeladze. Gut neurotoxin p-cresol induces brain-derived neurotrophic factor secretion and increases the expression of neurofilament subunits in PC-12 cells[J]. AIMS Neuroscience, 2022, 9(1): 12-23. doi: 10.3934/Neuroscience.2022002
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Abstract
Increased p-cresol levels reportedly alter brain dopamine metabolism and exacerbate neurological disorders in experimental animals. In contrast to toxic concentrations, low doses of p-cresol may have distinct effects on neuronal metabolism. However, the role of p-cresol in synapse remodeling, neurite outgrowth, and other anabolic processes in neurons remains elusive. We propose that low doses of p-cresol affect neuronal cell structural remodeling compared with the high concentration-mediated harmful effects. Thus, the effects of p-cresol on the secretion of brain-derived neurotrophic factor (BDNF) and neurofilament subunit expression were examined using rat pheochromocytoma cells (PC-12 cells). We observed that low doses of p-cresol potentiated nerve growth factor-induced differentiation via secretion of BDNF in cultured PC-12 cells. Opioidergic compounds modulated these p-cresol effects, which were reversed by oxytocin. We propose that this effect of p-cresol has an adaptive and compensatory character and can be attributed to the induction of oxidative stress. Accordingly, we hypothesize that low doses of p-cresol induce mild oxidative stress, stimulating BDNF release by activating redox-sensitive genes. Given that the intestinal microbiome is the primary source of endogenous p-cresol, the balance between gut microbiome strains (especially Clostridium species) and opioidergic compounds may directly influence neuroplasticity.
1.
Introduction
In population ecology, understanding how predators and primary producers influence nutrient flow relative to each other is important. Ecosystem interactions and predator-prey relationships are governed by predation and the delivery of resource processes. The identification of ecological factors that can alter or control dynamic behavior requires theoretical and experimental research. One way to study these questions is by means of experimental control, and another useful way is via mathematical modeling as well as computer simulations. Over decades of theoretical ecology and biomathematics development, mathematical modeling has become an indispensable tool for scientists in related fields to study ecosystems. Since Lotka [1] and Volterra [2], as cornerstones of theoretical ecology, published the first study of predator-prey dynamics, any species in nature can be a predator or prey, and due to its prevalence, it has become one of the most popular topics for researchers to study [3,4,5]. Besides, because biological resources are renewable and have the most unique development mechanisms, the over-utilization of biological resources and the destruction of the environment by humans will directly affect the balance of the ecosystem. Maintaining ecological balance and meeting humans material needs have attracted the most attention from researchers focused on the scientific management of renewable resource development [6,7,8].
Shelter serves as a defense strategy. It refers broadly to a series of behaviors by prey to avoid predators in order to increase their survival rate. The concept of sanctuary was first developed by Maynard-Smith [9] and Gause et al. [10], and its popularity has been very high, garnering widespread attention from many scholars [11,12,13,14,15]. Sih et al.[16] investigated the effects of prey refuge in a three-species model and concluded that the system's stability is strongly related to the refuge. Also, similar findings can be displayed in [17,18,19,20,21,22]. The two modes of refuge analyzed by Gonzalez-Olivares et al. [17] have diverse stability domains in terms of the parameter space. Qi et al.[21] ensure the stability of the system by varying the strength of the refuge.
Through reviewing a large amount of literature, we begin to consider [23,24] as a basis for the two prey and one predator species that will be modeled in this article. We assume that at a certain time t, the populations of the two prey and one predator are x1(t), x2(t), and y(t), respectively. Based on the above, we construct the following model:
Most species in nature, including humans, are influenced by fear. Fear may cause an abnormal state and behavior to arise. As usual, prey have an innate fear of predators. The ecology of fear is related to combining the optimal behavior of prey and predators with their population densities [25,26]. In view of reality, it is a fact that prey fear predators, which is seen as a psychological effect that can have a lasting impact on prey populations. This psychological influence is often easy to overlook, but it is necessary to consider it in the context of practical ecology [27]. Wang et al. [28] first considered the effect of the fear factor on the model and first proposed the fear of prey F(k,y). Afterwards, some researchers have investigated the effects of the fear effect and predator interferences in some three-dimensional systems as well as explored the generation of Hopf bifurcation conditions in the presence of a fear parameter as a bifurcation parameter [29,30,31,32]. Zanette et al. [33] observed that prey will reduce reproducing because of fear of being killed by predators, thus decreasing the risk of being killed after giving birth, which also leads directly to a decline in prey birth rates. According to the above discussion, our paper considers the different fears ki caused by predators for the two prey species.
In reality, when prey feel the crisis of being hunted, they will reproduce less and increase their survival rate. These conditions about the fear factor F(ki,y)(i=1,2) are listed as follows:
1)F(0,y)=1: prey production does not decrease when the prey does not fear the predator;
2)F(ki,0)=1: even though the prey will develop a fear of predators and there will be no predators, prey production will still not decline;
3)limki→∞F(ki,y)=0: when the prey's fear of the predator is very high, this will result in the prey production tending to zero;
4)limy→∞F(ki,y)=0: prey have a fear of predators, and when predator numbers are too large, this can also lead to prey production tending to zero;
5)∂F(ki,y)∂ki<0: the greater the prey's fear of predators, the less productive it will be;
6)∂F(ki,y)∂y<0: predators are inversely proportional to their prey.
For ease of analysis, we draw on Wang et al. [28] to consider the fear effect:
F(ki,y)=11+kiy(i=1,2),
(1.2)
obviously, F(ki,y)(i=1,2) in (1.2) satisfies conditions 1)–6). Based on the above conditions, this study will consider the effect of fear on system (1.1) to obtain system (1.3).
Notably, most biological parameters in much of the literature are fixed constants. However, in reality, the survival of species is full of unknowns, and all data are not always constant, which can lead to deviations from the ideal model with fixed parameters. In order to make the model more relevant and the results more accurate, we cannot just consider fixed parameters. Therefore, to make the study more convincing, it is necessary to target imprecise parameters. Professor Zadeh [34], who first proposed the fuzzy set theory, also argued that the application of fuzzy differential equations is a more accurate method for modeling biological dynamics in the absence of accurate data conditions [35]. Moreover, the first introduction of the idea of fuzzy derivatives came from Chang and Zadeh [36]. Further, Kaleva [37] studied the generalized fuzzy derivatives based on Hukuhara differentiability, the Zadeh extension principle, and the strong generalized differentiability concept. Bede et al. [38] employed the notion of strongly generalized differentiability to investigate fuzzy differential equations. Khastan and Nieto [39] solved the margin problem for fuzzy differential equations in their article. Motivated by the method of Pal [13] and Wang [23], we assume that the imprecise parameters ~r1, ~r2, ~a1, ~a2, ~c1, ~c2, ~e1, ~e1 and ˜d represent all triangular fuzzy numbers (the relevant theories of fuzzy sets are detailed in Appendix A), then the system (1.3) can be written as
The rest of the paper is shown below: In Section 2, we first prove the nonnegativity and boundedness of the system (1.7). Sections 3 and 4 discuss all possible equilibria and give conditions for the local asymptotic stability and global asymptotic stability of the equilibria. Immediately after that, in Section 5, we analyze the Hopf bifurcation by using the normal form theory. In Section 6, we numerically simulate the theoretical results of Sections 4 and 5. Finally, the article ends with detailed conclusions.
2.
Nonnegativity and boundedness
In this section, we give the following theorem to ensure the boundedness and nonnegativity of the solutions of the system (1.7).
Theorem 2.1.Provided that the initial values x1(0)>0, x2(0)>0, and y(0)>0, all solutions of system (1.7) are nonnegative.
Proof. It is not difficult to find that the right half of the system (1.7) fulfills the local Lipschitzian condition. Integrating both sides of the system (1.7) at the same time yields
If the solution curve starts at any internal point of R3+={(x1(t),x2(t),y(t))∈R3:x1(t)≥0, x2(t)≥0, y(t)≥0}, then x1(t), x2(t), and y(t) will always be nonnegative. □
Theorem 2.2.Assume that the initial values x1(0), x2(0), and y(0) are all greater than zero. The feasible region Ω is a positive invariant set of the system (1.7) defined by
It follows from the Descartes law of signs that Eq (3.4) has one and only one solution y∗ greater than zero if and only if g3>0, i.e., B1>x2K2B2+B3x1+q2E2. Substituting y∗ into the algebra expression on the right side of the first equation of the system (1.7) equals zero; furthermore, we obtain
Reusing the Descartes law of signs, we can assert that there exists at least one positive solution x∗1 of Eq (3.6) if and only if g4g5<0. And then we can deduce that
then the interior equilibrium P7(x∗1,x∗2,y∗) exists.□
4.
Stability analysis
In this section, the Jocabian matrix will be used to prove the local stability of all equilibria. Moreover, we prove the global stability of the internal equilibrium P7 by constructing a Lyapunov function.
4.1. Local stability
The Jocabian matrix for system (1.7) is given below:
The Routh-Hurwitz criterion shows that the internal equilibrium P7 is locally asymptotically stable; the following conditions need to be met: ψ1>0, ψ1ψ2>0, and ψ3>0.□
4.2. Global stability
This subsection studies the global asymptotic stability of interior equilibrium P7.
Theorem 4.2.If condition 4Γ1Γ2l1l2A2B2(1+k1y)(1+k2y) > (l1A3+l2B3)2 (i.e. 4Γ1Γ2l1l2(w1rα1R+w2rα1L)(w1rα2R+w2rα2L)(1+k1y)(1+k2y) > (l1(w1aα1R+w2aα1L)+l2(w1aα2R+w2aα2L))2) holds, then P7 is globally asymptotically stable.
Therefore, dVdt<0 if and only if 4Γ1Γ2l1l2A2B2(1+k1y)(1+k2y)>(l1A3+l2B3)2. □
5.
Hopf bifurcation
In this section, we will use the normal form theory introduced by Hassard et al.[40] and the central manifold theory [41] to study the Hopf bifurcation of the system (1.7). When the system (1.7) undergoes Hopf bifurcation, the corresponding characteristic equation must have a pair of conjugate pure imaginary roots, that is,
η1,2=±iω,i=√−1.
(5.1)
Consider the parameter k1 as a bifurcation parameter. When the value of parameter k1 changes near the critical point kΞ1 of Hopf bifurcation, the pure imaginary roots ±iω will become a complex eigenvalue η=ρ+i˜ω. Substituting η=ρ+i˜ω into Eq (4.12), we need to separate the imaginary and real parts to get
ρ3+ψ3+ρψ2+ρ2ψ1−3ρ˜ω2−ψ1˜ω2=0,
(5.2)
3ρ2˜ω+ψ2˜ω+2ρψ1˜ω−˜ω3=0.
(5.3)
By simplifying Eqs (5.2) and (5.3), we obtain
ψ3−8ρ3−2ρψ2−8ρ2ψ1−ψ1ψ2−2ρψ21=0,
(5.4)
at k1=kΞ1, taking the derivative of Eq (5.4) over k1 yields
If it satisfies dρdk1|k1=kΞ1≠0, the system (1.7) will generate Hopf bifurcation, which indicates that when parameter k1 crosses the bifurcation critical point kΞ1, the population state evolves from stable equilibrium to periodic oscillation over time.
When the system (1.7) undergoes Hopf bifurcation at k1=kΞ1, the final decision condition is also met. Considering that the characteristic roots of Eq (4.12) are η1,2=±iω and η3=−ψ1, in order to obtain this condition, we introduce
z1=x1−x∗1,z2=x2−x∗2,z3=y−y∗.
(5.6)
Substituting (5.6) into the system (1.7) and separating the linear and nonlinear parts, it can be obtained that
where O((|z1|+|z2|+|z3|)4) is a fourth-order polynomial function about variables (|z1|,|z2|,|z3|), while tj1j2j3, nj1j2j3, and lj1j2j3 can be obtained through calculation:
Correspondingly, the dynamic properties of the system are limited to the central flow Wc(0,0,0), and in conjunction with Eq (5.14), system (5.12) can be simplified as
where subscripts y1 and y2 indicate partial derivatives for the first and second variable, respectively. Based on Eq (5.18), it can be obtained that Uy1=0, Uy2≠0, Ny1≠0, Ny2=0, and Uy2Ny1≠0. In addition, it ensures that the system (5.18) has pure virtual feature roots ±i√|Uy2Ny1|. Thus, it can be determined that system (1.7) produces Hopf bifurcation; the direction of the bifurcation is determined by the following equation:
QkΞ1=116ω(ℓ3+ℓ5+ȷ5+ȷ7)+116ω(ℓ1ℓ3−ȷ2ȷ3−ȷ1ȷ2−ȷ1ℓ1).
(5.21)
Theorem 5.1.If dρdk1|k1=kΞ1≠0, then system (1.7) will generate Hopf bifurcation at interior equilibrium P7. In addition, when dρdk1|k1=kΞ1<0, if QkΞ1<0 and 0<k1−kΞ1≪1, then system (1.7) will generate supercritical Hopf bifurcation and form a stable periodic orbit, or if QkΞ1>0 and 0<k1−kΞ1≪1, then system (1.7) will generate subcritical Hopf bifurcation and form a stable periodic orbit.
6.
Numerical simulations
In this section, we first discussed equilibria P1 to P7 of system (1.7) with distinct values of α, w1, and w2. Consider the parameter values as follows: ~r1=(2.8,3,3.2), ~r2=(2.8,3,3.2), ~c1=(0.1,0.2,0.3), ~c2=(0.5,0.6,0.7), ~a1=(0.1,0.2,0.3), ~a2=(0.2,0.3,0.4), ~e1=(0.2,0.3,0.4), ~e2=(0.3,0.4,0.5), and ˜d=(0.1,0.2,0.3). Tables 2–8 showed that the trivial equilibrium P1 retained constant at (0, 0, 0), the values of prey x1, prey x2, and predator y always maintained at 0; the values of prey x1 in P2 and prey x2 in P3 severally decreased with increasing w1 under the same α; the values of prey x1 and predator y in P4 increased with increasing w1, and for P5 the value of prey x2 and predator y rose with growing w1; the values of prey x1 and x2 in P6 decreased with growing w1; and for the same α, considering interior equilibrium P7, the values of prey x1, prey x2, and predator y decreased with growing w1.
Table 2.
The trivial equilibrium P1 for k1=0.1, k2=0.7, q1=0.7, q2=0.5, q3=0.7, E1=0.3, E2=0.2, E3=0.2, K1=5, K2=5, m1=0.9, m2=0.3.
Considering four sets of different initial values, it could be seen from Figure 1 that different orbits eventually converged to the same value, which concluded that the interior equilibrium of the system (1.7) fulfills the character of globally asymptotical stability. Figure 2 plotted the bifurcation graph of system (1.7) with the horizontal coordinates k1, and the Hopf bifurcation of the system occurred with k1 taking values in the range of 0.01≤k1≤0.7. When 0.01≤k1<0.384, the system oscillates periodically, while it maintains a stable steady-state when 0.384<k1≤0.7. Therefore, based on Figure 2, it could be concluded that the fear of prey x1 for predator y affected the stability of the system. We further observed that as k1 increased, the prey x1 density continued to decrease while the predator y density kept increasing. Thus, the result also suggested that greater fear of predators had a negative impact on prey populations while having a positive impact on predator populations. Correspondingly, Figures 3 and 4 showed the waveform plots and phase diagram at k1=0.1 and k1=0.7, respectively.
Figure 1.
Global stability of the internal equilibrium P7 = (5.665, 1.668, 2.047) of system (1.7) is given by the following parameter values: α=1, w1+w2=1, A1=2.0, A2=2.0, B1=2.0, B2=2.0, k1=0.2, k2=0.1, q1=0.4, q2=0.4, q3=0.2, E1=0.2, E2=0.2, E3=0.2, A3=0.1, B3=0.1, A4=0.3, B4=0.6, K1=10, K2=10, m1=0.4, m2=0.4, C1=0.1, C2=0.2, C3=0.5.
In addition, Figure 5 also plots the bifurcation graph with changing m1. As can be seen in Figure 5, m1 took values from 0.3 to 1, in which the system also underwent a Hopf bifurcation. When the value m1 ranged from 0.3 to 0.657, the system (1.7) was stable; nevertheless, it would become unstable at 0.657<m1≤1. Correspondingly, Figures 6 and 7 showed the waveform plots and phase diagram at m1=0.6 and m1=0.9, respectively.
Figure 5.
Hopf bifurcation occurs as a bifurcation parameter of system (1.7) parameter m1, and the remaining parameters take the following values: α=1, w1+w2=1, A1=2.0, A2=2.0, B1=2.0, B2=2.0, k1=0.1, k2=0.4, q1=0.7, q2=0.4, q3=0.2, E1=0.2, E2=0.2, E3=0.3, A3=0.1, B3=0.1, A4=0.3, B4=0.6, K1=10, K2=70, m2=0.4, C1=0.1, C2=0.2, C3=0.5.
Further, we find an interesting dynamic phenomenon through some numerical simulations. System (1.7) appears as a chaotic phenomenon, as shown in Figure 8.
Figure 8.
Waveform plots and phase diagram of chaotic phenomena with the following parameter values: α=1, w1+w2=1, A1=2.0, A2=2.0, B1=3.0, B2=3.0, k1=0.2, k2=0.5, q1=0.6, q2=0.4, q3=0.2, E1=0.2, E2=0.3, E3=0.2, A3=0.2, B3=0.3, A4=0.3, B4=0.6, K1=10, K2=70, m1=0.9, m2=0.3, C1=0.1, C2=0.2, C3=0.5.
In this work, we develop a model of one-predator and two-prey interactions in a fuzzy environment, examine the effects of fear and prey refuge on the system, and provide insight into the dynamic complexity. The proofs of the theoretical parts of this paper are based on system (1.7). It has been proven that all equilibria in system (1.7) are locally asymptotically stable, and interior equilibrium P7 is also globally asymptotically stable. We have been further concerned about the appearance and direction of Hopf bifurcation. With the support of theoretical research, our numerical simulations have been able to display a wealth of charts and graphs.
First of all, different equilibria are displayed from Tables 2–8 with different α,w1,w2, respectively. Throughout Figure 1, we have verified the global asymptotical stability of interior equilibrium P7, and find that the system is from unstable to stable with the increase of fear k1, which demonstrates that the fear effect may be an important factor influencing the stability of the system (see Figures 2–4). Furthermore, it has also been observed that an increase in prey refuge m1 leads to oscillatory phenomena (see Figures 5–7). Finally, through studying the Hopf bifurcation, we have discovered some interesting biological phenomena, namely that system (1.7) appears to be in a chaotic state (see Figure 8).
Author contributions
Xuyang Cao: Conceptualization, Investigation, Methodology, Validation, Writing-original draft, Formal analysis, Software; Qinglong Wang: Conceptualization, Methodology, Formal analysis, Writing-review and editing, Supervision; Jie Liu: Validation, Visualization, Data curation.
Use of AI tools declaration
The authors declare they have not used Artificial Intelligence (AI) tools in the creation of this article.
Acknowledgments
The authors thank the editor and referees for their careful reading and valuable comments.
The work is supported by the Natural Science Foundation of Hubei Province (No. 2023AFB1095) and the National Natural Science Foundation of China (No. 12101211) and the Program for Innovative Research Team of the Higher Education Institution of Hubei Province (No. T201812) and the Teaching Research Project of Education Department of Hubei Province (No. 2022367) and the Graduate Education Innovation Project of Hubei Minzu University (Nos. MYK2024071, MYK2023042).
Conflict of interest
The authors declare that there are no conflicts of interest regarding the publication of this paper.
Appendix A
Definition 1.[34] Fuzzy set: A fuzzy set ˜ℏ in a universe of discourse S is denoted by the set of pairs
˜ℏ={(s,μ˜ℏ(s)):s∈S},
where the mapping μ˜ℏ:S→[0,1] is the membership function of the fuzzy set ˜ℏ and μ˜ℏ is the membership value or degree of membership of s∈S in the fuzzy set ˜ℏ.
Definition 2.[42] α-cut of fuzzy set: For any α∈(0,1], the α-cut of fuzzy set ˜ℏ defined by ℏα={s:μ˜ℏ(s))≥α} is a crisp set. For α=0 the support of ˜ℏ is defined as ℏ0=Supp(˜ℏ)=¯{s∈R,μ˜ℏ(s)>0}.
Definition 3.[43] Fuzzy number: A fuzzy number satisfying the property S=R is called a convex fuzzy set.
Definition 4.[44] Triangular fuzzy number: A triangular fuzzy number (TFN) ˜ℏ≡(b1,b2,b3) represent fuzzy set of the real line R satisfying the property that the membership function μ˜ℏ:R→[0,1] can be espressed by
Hence, the α-cut of triangular fuzzy number meets boundedness and encapsulation on [ℏL(α),ℏR(α)], in which ℏL(α)=infs:μ˜ℏ(s)≥α=b1+α(b2−b1) and ℏR(α)=sup{s:μ˜ℏ(s)≥α}=b3+α(b3−b2).
Lemma 1.[45] In weighted sum method, wj stands for the weight of jth objective. wjgj represent a utility function for jth objective, and the total utility function π is represented by
π=l∑jwjgj,j=1,2,⋯,l,
where wj>0 and ∑ljwj=1 are satisfied.
Acknowledgments
This research was supported by the Shota Rustaveli National Science Foundation of Georgia (grant # FR -19-3114).
Conflict of interests
The authors declare that there are no conflicts of interest.
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Yuan Tian, Hua Guo, Wenyu Shen, Xinrui Yan, Jie Zheng, Kaibiao Sun,
Dynamic analysis and validation of a prey-predator model based on fish harvesting and discontinuous prey refuge effect in uncertain environments,
2025,
33,
2688-1594,
973,
10.3934/era.2025044
Interspecific competition between prey x1 and prey x2
c1,c2
Predation coefficients for prey x1 and prey x2
m1,m2
Refuge rates of prey x1 and prey x2
e1,e2
Conversion factors for prey x1 and prey x2
q1,q2,q3
Captureability factors for prey x1, prey x2 and predator y
E1,E2,E3
Harvesting efforts for prey x1, prey x2 and predator y
d
Predator y mortality rate
w1
w2
P1 at α=0
P1 at α=0.3
P1 at α=0.6
P1 at α=0.9
0
1
(0,0,0)
(0,0,0)
(0,0,0)
(0,0,0)
0.2
0.8
(0,0,0)
(0,0,0)
(0,0,0)
(0,0,0)
0.4
0.6
(0,0,0)
(0,0,0)
(0,0,0)
(0,0,0)
0.6
0.4
(0,0,0)
(0,0,0)
(0,0,0)
(0,0,0)
0.8
0.2
(0,0,0)
(0,0,0)
(0,0,0)
(0,0,0)
1
0
(0,0,0)
(0,0,0)
(0,0,0)
(0,0,0)
w1
w2
P2 at α=0
P2 at α=0.3
P2 at α=0.6
P2 at α=0.9
0
1
(5.3393,0,0)
(5.1224,0,0)
(4.9144,0,0)
(4.7148,0,0)
0.2
0.8
(5.0521,0,0)
(4.9280,0,0)
(4.8069,0,0)
(4.6888,0,0)
0.4
0.6
(4.7804,0,0)
(4.7409,0,0)
(4.7017,0,0)
(4.6629,0,0)
0.6
0.4
(4.5230,0,0)
(4.5608,0,0)
(4.5988,0,0)
(4.6372,0,0)
0.8
0.2
(4.2788,0,0)
(4.3872,0,0)
(4.4980,0,0)
(4.6116,0,0)
1
0
(4.0469,0,0)
(4.2197,0,0)
(4.3994,0,0)
(4.5861,0,0)
w1
w2
P3 at α=0
P3 at α=0.3
P3 at α=0.6
P3 at α=0.9
0
1
(0,5.5357,0)
(0,5.3147,0)
(0,5.1027,0)
(0,4.8993,0)
0.2
0.8
(0,5.2431,0)
(0,5.1166,0)
(0,4.9932,0)
(0,4.8728,0)
0.4
0.6
(0,4.9662,0)
(0,4.9260,0)
(0,4.8861,0)
(0,4.8465,0)
0.6
0.4
(0,4.7039,0)
(0,4.7424,0)
(0,4.7812,0)
(0,4.8202,0)
0.8
0.2
(0,4.4551,0)
(0,4.5655,0)
(0,4.6785,0)
(0,4.7942,0)
1
0
(0,4.2187,0)
(0,4.3949,0)
(0,4.5779,0)
(0,4.7682,0)
w1
w2
P4 at α=0
P4 at α=0.3
P4 at α=0.6
P4 at α=0.9
0
1
(0.4800,0,0.1496)
(0.5838,0,0.1767)
(0.7059,0,0.2040)
(0.8516,0,0.2317)
0.2
0.8
(0.6222,0,0.1858)
(0.6971,0,0.2022)
(0.7802,0,0.2188)
(0.8732,0,0.2354)
0.4
0.6
(0.8000,0,0.2224)
(0.8306,0,0.2280)
(0.8623,0,0.2335)
(0.8954,0,0.2391)
0.6
0.4
(1.0286,0,0.2595)
(0.9902,0,0.2539)
(0.9534,0,0.2484)
(0.9181,0,0.2428)
0.8
0.2
(1.3333,0,0.2969)
(1.1845,0,0.2801)
(1.0551,0,0.2633)
(0.9415,0,0.2465)
1
0
(1.7600,0,0.3343)
(1.4261,0,0.3062)
(1.1692,0,0.2782)
(0.9655,0,0.2502)
w1
w2
P5 at α=0
P5 at α=0.3
P5 at α=0.6
P5 at α=0.9
0
1
(0,0.1920,0.3638)
(0,0.2298,0.3833)
(0,0.2727,0.4029)
(0,0.3220,0.4226)
0.2
0.8
(0,0.2435,0.3899)
(0,0.2697,0.4016)
(0,0.2981,0.4134)
(0,0.3291,0.4252)
0.4
0.6
(0,0.3048,0.4160)
(0,0.3150,0.4199)
(0,0.3255,0.4239)
(0,0.3363,0.4278)
0.6
0.4
(0,0.3789,0.4422)
(0,0.3668,0.4383)
(0,0.3551,0.4343)
(0,0.3437,0.4304)
0.8
0.2
(0,0.4706,0.4683)
(0,0.4268,0.4566)
(0,0.3872,0.4448)
(0,0.3513,0.4330)
1
0
(0,0.5867,0.4942)
(0,0.4970,0.4748)
(0,0.4222,0.4553)
(0,0.3590,0.4357)
w1
w2
P6 at α=0
P6 at α=0.3
P6 at α=0.6
P6 at α=0.9
0
1
(4.9826,3.8455,0)
(4.6419,3.5356,0)
(4.3385,3.2568,0)
(4.0671,3.0043,0)
0.2
0.8
(4.5369,3.4395,0)
(4.3577,3.2745,0)
(4.1901,3.1191,0)
(4.0331,2.9724,0)
0.4
0.6
(4.1543,3.0858,0)
(4.1016,3.0366,0)
(4.0500,2.9883,0)
(3.9995,2.9408,0)
0.6
0.4
(3.8232,2.7740,0)
(3.8700,2.8184,0)
(3.9177,2.8636,0)
(3.9665,2.9097,0)
0.8
0.2
(3.5351,2.4958,0)
(3.6599,2.6172,0)
(3.7926,2.7447,0)
(3.9338,2.8789,0)
1
0
(3.2834,2.2448,0)
(3.4690,2.4307,0)
(3.6742,2.6311,0)
(3.9017,2.8485,0)
w1
w2
P7 at α=0
P7 at α=0.3
0
1
(1.7240,3.6276,1.1986)
(1.5910,3.4597,1.0271)
0.2
0.8
(1.6168,3.5123,1.0583)
(1.5042,3.3223,0.9139)
0.4
0.6
(1.5417,3.4296,0.9337)
(1.4211,3.1793,0.8023)
0.6
0.4
(1.4309,3.2886,0.7930)
(1.3913,3.1413,0.7269)
0.8
0.2
(1.4291,3.2451,0.6884)
(1.3195,3.0467,0.6350)
1
0
(1.3764,2.9890,0.5245)
(1.2797,2.9766,0.5529)
w1
w2
P7 at α=0.6
P7 at α=0.9
0
1
(1.7066,3.7444,1.0113)
(1.5325,3.4231,0.7999)
0.2
0.8
(1.5291,3.4203,0.8678)
(1.4899,3.3773,0.7762)
0.4
0.6
(1.5273,3.4215,0.8306)
(1.4789,3.3677,0.7638)
0.6
0.4
(1.4894,3.3737,0.7779)
(1.4685,3.3587,0.7516)
0.8
0.2
(1.4594,3.3320,0.7279)
(1.4590,3.3504,0.7397)
1
0
(1.4328,3.2552,0.6687)
(1.4124,3.2841,0.7105)
Figure 1. Microscope images of PC-12 cell culture (Jenco, Inverted light microscope; total magnification 400×): a) PC-12 cells without treatment at day 8; b) PC-12 cells treated with 1 µM p-cresol at day 8; c) PC-12 cells treated with NGF for 3 days, no pretreatment with 1 µM p-cresol; d) PC-12 cells treated with NGF for 3 days after 5-day pretreatment with 1 µM p-cresol. NGF, nerve growth factor. e) Quantification of the neurite length (µm) using Image J software (https://imagej.net/ImageJ). The values represent averages SEM from three independent cultures. The significance level was set at *p < 0.05, versus the NGF treated PC-12 cells
Figure 2. Analysis of the neurofilament (NF) heavy (H) and medium (M)-chain content in PC-12 cells. The cell lysates were loaded onto each well for immunoblotting, resolved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), transferred to nitrocellulose membranes, and probed with NF-H/M antibodies. Bands from three experiments (n = 3) were quantified using Image J software (https://imagej.net/ImageJ). and plotted as a bar graph. Quantitative values represent mean normalized to total protein. The significance level was set at *p < 0.05, versus the NGF treated PC-12 cells. Data are derived from three independent experiments and are expressed as the relative change from the mean ± standard error of triplicate experiments' mean (SEM)
Figure 3. Analysis of the BDNF content in the incubation medium. The incubation period—24 hours. The significance level was set at *p < 0.05, displays versus the untreated PC-12 cells (control). Data are derived from three independent experiments (n = 3) and are expressed as the relative change from the mean ± standard error of triplicate experiments' mean (SEM). BDNF, brain-derived neurotrophic factor
Figure 4. Analysis of the BDNF content in the incubation medium. The incubation period—48 hours. The significance level was set at *p < 0.05, versus the untreated PC-12 cells (control), and #p < 0.05, versus p-cresol 1 µM treated group. Data are derived from three independent experiments (n = 3) and are expressed as the relative change from the mean ± standard error of triplicate experiments' mean (SEM)