Abstract
The solvothermal reactions of C3-symmerical 4,4,4-nitrilotribenzoic acid and MnCl2 afforded a novel Mn(II) compound, and its formula is [Mn3(TCA)2(e-urea)2(DMA)2]n·n(urea)·n(DMA) (1, H3TCA=4,4,4-nitrilotribenzoic acid, e-urea=2-imidazaolidone, DMA=N,N-dimethylacetamide). Magnetic property investigation of 1 indicates weak antiferromagnetic mutual effects exist between neighboring Mn(II) ions. Serial biological tests were adopted to discover new compound activity. Firstly, the enzyme-linked immunosorbent assay (ELISA) was conducted to measure the content of inflammatory cytokines released into the synovial fluid. In addition, we also studied adenosine 5-monophosphate- (AMP-) activated protein kinase (AMPK) inflammatory signaling pathway activation through real-time reverse transcription-polymerase chain reaction (RT-PCR).
1. Introduction
Osteoarthritis, also known as degenerative joint disease, is the most common of all arthritis. Its incidence increases exponentially with age [1]. Among people over 65, 80-90% will suffer from arthritis. Various factors can induce secondary arthritis, such as obesity, hemochromatosis, and melasma. Nowadays, more and more researches are aimed at developing new candidates for the foot and ankle arthritis therapy [2].
Metal-organic frameworks (MOFs) which undergone an explosive development over the past few years have attracted considerable attention largely attributed to their potentials in luminescence, gas storage and separation, magnetism, heterogeneous catalysis, and so on [3–7]. From the reports, it can be found that the most successful class of organic ligands for the syntheses of functional MOFs is the multicarboxylate ligands [8–11]. The main advantages of the polycarboxylate ligands in the syntheses of MOFs are as follows: (i) the polycarboxylate ligands have good coordination capacities to transition metal ions; (ii) the carboxylate groups of ligand can utilize diversified coordination patterns based on the geometrical requirement of central metal ions; (iii) aggregates of metal-carboxylate group often referred to as the second building subunits (SBUs) are easily formed through linkage of carboxylate groups together with metal ions; and (iv) the polycarboxylate ligands can adopt different anionic forms according to the pH of reaction system [12–14]. On the other hand, much efforts have been devoted to the design and preparation of transition metal MOF-based materials because these metals are physiologically essential metal elements that function in endogenous oxidative DNA damage associated with aging and cancer. Metal ions that are essential for cells may become toxic at high concentrations because they can replace other important metals in enzymes. Besides, MOFs can interact with DNA through noncovalent interactions, including intercalation and groove binding for large molecules and slot external electrostatic binding for cations [15]. Many MOFs with interesting bioactivity have been studied and reported. For instance, Chandra et al. reported a 3D Zn-MOF prepared using the mixed-ligand approach, and the biological studies suggest that this MOF is antibacterial as well as a promising anticancer agent [16]; Jaros et al. discovered that the Ag-MOF has revealed distinctive bioactive features, namely, (i) a remarkable antiviral activity against herpes simplex virus type 1 (HSV-1) and adenovirus 36 (Ad-36), (ii) a significant antibacterial activity against clinically important bacteria (Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa), and (iii) a selective cytotoxicity against HeLa (human cervix carcinoma) cell line [17]. In viewing of these advantages of MOFs, a C3-symmetric ligand of 4,4,4-nitrilotribenzoic acid was chosen in this study, which has three carboxylate groups and features a triphenylamine core, to react with MnCl2 under solvolthermal conditions. Successfully, a novel Mn(II) compound, namely, [Mn3(TCA)2(e-urea)2(DMA)2]n·n(urea)·n(DMA) (1, H3TCA=4,4,4-nitrilotribenzoic acid, e-urea=2-imidazaolidone, DMA=N,N-dimethylacetamide, Scheme 1), was prepared and further structurally shaped by the single crystal X-ray diffraction analysis, elemental analysis, powder X-ray diffraction analysis, and thermogravimetric analysis. Its structure is a 2D layer which is on the accordance with linear trinuclear [Mn3(COO)6] cluster-based subunits. We discovered thermal behavior and magnetic property of 1 as well. Furthermore, new complex’s treatment activity on the foot along with ankle arthritis was assessed, and we detected its mechanism.
2. Experimental
2.1. Materials and Instrumentation
Used chemicals are of analytic grade and commercially available from Jinan Henghua Company. Through the utilization of an elemental Vario EL III analyzer, elemental analyses including C and H along with N were completed. The powder diffractometer of PANalytical X’Pert Pro was adopted for PXRD analysis for 1 utilizing Cu/Kα radiation (with λ of 1.54056 Å). The TGA data for 1 were completed through NETSCHZ STA-449C application under flow of nitrogen at 10°C/min heating rate in 30 to 800°C temperature. The data of single crystal structure was gathered through the graphite-monochromated Mo-Kα radiation (with λ of 0.71073 Å) in the environment of room temperature via exploiting Mercury CCD diffractometer.
2.2. Synthesis of Compound [Mn3(TCA)2(e-urea)2(DMA)2]n·n(urea)·n(DMA)(1)
The mixture formed by 0.1 mmol of MnCl2, 0.05 mmol of H3TCA, 0.2 mmol of e-urea (0.2 mmol), and 2 mL of DMA (2 mL) along with 1 mL water was sealed to a small glass vial (20 mL). The obtained mixture was heated at 110°C temperature lasting three days. After the mixture was naturally cooled to the environmental temperature gradually, compound’s colorless block crystals of 1 were achieved with yield of 38% in the light of H3TCA. Elemental analysis calcd. for C70H84Mn3N14O20 (1606.33): C, 52.29; H, 5.23; N, 12.20%. Found: C, 52.04; H, 5.45; N, 11.96%. IR (KBr, cm−1): 3398 s, 1608 s, 1488 m, 1411s, 1352 m, 1281 m, 1248 w, 1212 s, 1172 s, 1101 s, 1070 m, 1032 s, 947 s, 838 w, 865 s, 829 w, 811 s, 762 s, 721 m, 657 m, 637 m.
2.3. X-Ray Crystallography
With the assistance of direct means, compound 1’s crystal architecture can be completed with ShelXS structure solution program, and then, ShelXL refinement package is applied for refinement via full-matrix least square technique on the basis of F2 [18]. Anistropical refinement of total nonhydrogen atoms was conducted through anisotropic displacement parameters together with riding model. The data of crystallography and refinements of 1 are revealed in Table 1. The chosen bond angles (°) and lengths (Å) around central metal ion are exhibited in Table S1.
2.4. ELISA Assay
To determine the content of inflammatory cytokines released into synovial fluid after complex treatment, the application of ELISA detection kit was completed in this work. This study was accomplished which completely adheres to the instructions with limited adjustments. Shortly, fifty BALB/c mice utilized were obtained from the Nanjing University (Nanjing, China). The overall operation of this experiment had gained approval from Animal Ethics Committee of Nanjing University (Nanjing, China). The LPS was injected into mice to lead to foot and ankle arthritis animal model, and then, animal was disposed by compound at the concentration of 1, 2, and 5 mg/kg. Finally, synovial fluid was gathered, and inflammatory cytokines content was measured at least three times.
2.5. Real-Time RT-CPR
To evaluate the activation of AMPK inflammatory signaling pathway in the ankle tissue, real-time RT-PCR was performed in this present research. This experiment was conducted strictly complying with guidance of the instructions accompanied by some adjustment. In a word, fifty BALB/c mice utilized were obtained from the Nanjing University (Nanjing, China). The overall operation of this experiment had gained approval from Animal Ethics Committee of Nanjing University (Nanjing, China). The LPS was injected into mice to lead to foot and ankle arthritis animal model, and then, animal was disposed by compound at the concentration of 1, 2, and 5 mg/kg. In the next step, ankle tissue was collected from different groups, and total RNA inside brain was extracted through TRIzol reagent. The subsequent step is to measure its quality and quantity. Afterwards, it completed the reverse transcript into cDNA. Lastly, real-time RT-PCR was exercised, and relative expression of AMPK inflammatory signaling pathway in the ankle tissue was determined with gapdh utilized as the control gene.
3. Results and Discussion
3.1. Crystal Structure of Compound 1
The X-ray crystallographic analysis revealed that the structure of 1 crystallizes in triclinic P-1 space group and displays a two-dimensional layered construction on the accordance with linear trinuclear [Mn3(COO)6] clusters. Asymmetric unit of 1 includes 1.5 Mn(II) ions, 1 TCA3- ligand, 1 coordinated e-urea molecule, 1 coordinated DMA molecule, and 1 lattice e-urea molecule together with 1 lattice DMA molecule. We can observe from Figure 1(a) that coordination geometries of two independent Mn(II) centers can be described as OC-6 (octahedral) geometries as calculated via the SHAPE software, and the calculated parameter was 0.086 for Mn1 and 3.750 for Mn2 [19]. The octahedron of Mn1 center is defined by six carboxylate oxygen atoms offered by six different TCA3- ligands, and octahedron of Mn2 center is determined by four carboxylate oxygen atoms from three distinct TCA3- ligands and two oxygen atoms from one e-urea molecule along with one DMA molecule. In the structure of 1, the H3TCA ligand is fully deprotonated, and each TCA3- ligand acts as hexadentate ligand linking with six distinct Mn(II) centers in a (κ1-κ1)-(κ1-κ1)-(κ2-μ2)-μ6 pattern (Fig. S1). The scope of bond lengths around Mn(II) centers is 2.1108 (17)-2.3283 (17) Å, while bond angles of O-Mn(II)-O are in the range of 57.56 (6) to 180.00° that are listed in Table S1. Four bis-monodentate carboxylate groups and two chelating-bridged carboxylate groups linked one Mn1 center and two symmetry-related Mn2 centers together, leading to the formation of a linear trinuclear [Mn3(COO)6] cluster (Figure 2(b)), where neighboring Mn…Mn distance is 3.62 Å. Further, TCA3- ligands act as connectors linking these linear trinuclear into a 2D layer (Figure 1(c)). If the TCA3- ligands and linear trinuclear [Mn3(COO)6] clusters are able to be regarded as 3- and 6-linked nodes separately, such two-dimensional layer is able to be topologically simplified into a (3,6)-linked kgd net with the point symbol of {43}2{46.66.83} (Figure 1(d)). Finally, these adjacent 2D layers further stacked in -AA- mode induced by the weak van der Waals forces, finally affording a 3D supramolecular framework (Figure 1(e)). The free DMA and e-urea molecules filled in the free voids of this 3D supramolecular framework.
(a)
(b)
(c)
(d)
(e)
(a)
(b)
3.2. Powder X-Ray Diffraction Patterns (PXRD) and Thermogravimetric Analyses (TGA)
With the aid of PXRD characterization, it is found that as-synthesized samples’ mode has a good match with mode in line with single crystal diffraction information, demonstrating good purity of the bulk products (Figure 2(a)).
In addition, thermogravimetric analysis experiment was adopted to discover thermal behavior of 1 under nitrogen atmosphere as well (Figure 2(b)). The weight loss (43.23%) in the first step happened under the temperature area of 105 to 198°C indicating the complete removal of the free and coordinated solvent molecules (DMA and e-urea) with the calculated value of 43.13%. The skeleton of the desolvated sample is able to be stable up to 295°C, after which we can notice sharp weight loss that was caused by organic ligand decomposition. Final decomposition products with a weight of 13.16% indicate the formation of MnO (calcd: 13.25%).
3.3. Magnetic Property of 1
The magnetic property of 1 was discovered at an applied magnetic area of one thousand Oe, and temperature-dependent χMT along with χM-1 data is revealed in Figure 3. According to plot of χM-1 versus T, it is obvious that the susceptibility data can be well fitted into a line in the temperature scope of 2-300 K, which obeys Curie-Weiss law and gives a Weiss constant K and a Curie constant cm3·K·mol−1. From the plot of χMT versus T, we found that the χMT value at 300 K is 12.54 cm3·K·mol−1 per Mn3 unit, which is a little lower than that of three magnetically isolated Mn(II) ions (, ) with the spin-only value of 13.12 cm3·K·mol−1, and further decreased smoothly to 4.48 cm3·K·mol−1 upon cooling to 2 K. Large negative value of Weiss constant and temperature-dependent χMT behavior indicate that the dominant mutual effects between neighboring Mn(II) ions in 1 are antiferromagnetic. To further estimate the magnetic coupling mutual effects between nearby Mn(II) centers, susceptibility data of 1 were simulated using a trinuclear model by assuming the same interactions of Mn1-Mn2 and Mn1-Mn2a. The magnetic coupling interactions between the Mn(II) centers is regulated by the Heisenberg Spin Hamiltonian equation: , of which J is the exchange interactions between nearby Mn(II) ions and and , together with , are typical spin vectors [20]. The least squares curve fitting by MAGPACK program causes below outcomes: and [21]. Negative J constant implies that antiferromagnetic mutual effects exist between the neighboring Mn(II) ions in the linear trinuclear Mn3 cluster.
3.4. Compound Distinctly Decreased Inflammatory Cytokines Releasing into Synovial Fluid
After the synthesis of the novel complex, its biological activity was firstly discovered. There was usually a distinctly improved degree of inflammatory cytokines released into synovial fluid. Thus, we adopted the ELISA detection kit to measure the content of inflammatory cytokines released into synovial fluid after indicated compound treatment. We can notice from Figure 4 that there was a distinctly improved degree of inflammatory cytokines in the model team, compared with normal team, accompanied by . After novel complex treatment, content of inflammatory cytokines released into synovial fluid was markedly decreased. New compound’s restrain exhibited a dose-dependent manner.
3.5. Compound Markedly Restrained Activation of AMPK Inflammatory Signaling Pathway
It is proved in the above study that the complex has excellent application values on the foot and ankle arthritis treatment by inhibiting the inflammatory cytokines releasing. As the AMPK inflammatory signaling pathway regulated inflammatory cytokines releasing, therefore we also determined activation of AMPK inflammatory signaling pathway through real-time RT-PCR assay. Outcomes in Figure 5 revealed that activation of AMPK inflammatory signaling in model team was much higher than that of the control team. Under new compound disposal, activation of AMPK inflammatory signaling was obviously decreased in a dose relationship.
4. Conclusion
In summary, we successfully synthesized a novel trinuclear cluster-based Mn(II) compound with a 2D layered structure. This 2D layer is able to be topologically simplified into a (3,6)-linked kgd net by viewing the TCA3- ligands together with trinuclear [Mn3(COO)6] cluster as 3- and 6-linked nodes, respectively. Moreover, such compound exhibits antiferromagnetic property. Outcomes of ELISA assay indicated that the complex could distinctly decrease inflammatory cytokines releasing into the synovial fluid. Real-time RT-PCR information suggested that activation of AMPK inflammatory signaling pathway was restrained by new complex as well. At last, we reached this conclusion; complex could be an excellent candidate for the foot and ankle arthritis by reducing the over inflammatory response.
Data Availability
Selected bond lengths (Å) and angles (°) for 1 are found in Table S1. The coordination mode of TCA3- ligand in 1 is found in Fig. S1. The information could be found in the supporting information file.
Conflicts of Interest
The authors declare that there is no conflict of interest regarding the publication of this paper.
Supplementary Materials
Table S1: selected bond lengths (Å) and angles (°) for 1. Fig. S1: the coordination mode of TCA3- ligand in 1. (Supplementary Materials)