Analysis of full and empty ratio of EV71 virus by using capillary zone electrophoresis.

Hand, foot, and mouth disease is a serious public health problem, and the main pathogen is enterovirus 71 (EV71). Its capsid assembly mechanism including capsid protein processing has been widely studied. Full and empty capsids have different immunological efficacy. Therefore, tracking full/empty capsid ratio throughout the EV71 production process is important to ensure consistent product quality and proper dosing response. The analysis of full/empty capsid ratio of intact virus has been widely reported as well. A variety of techniques have been employed to evaluate the full/empty capsid ratios. However, there has not been a rapid, reproducible, and robust assay to determine the full/empty capsid ratios of final and in-process products. In this study, a novel assay based on capillary zone electrophoresis was established. The separation of full and empty species could be achieved within 10 min and the ratio of peak areas was used to calculate the full/empty capsid ratio directly. The results showed good reproducibility and linearity for the determination of full/empty capsid ratios.

Letter to the Editor concerning the article "Clinical outcomes of chondroblastoma treated using synthetic bone substitute: risk factors for developing radiographic joint degeneration".

The purpose of this letter to the Editor is to report some shortcomings in the statistical analysis and variable grouping in the recent publication of the article "Clinical outcomes of chondroblastoma treated using synthetic bone substitute: risk factors for developing radiographic joint degeneration," and to further explore some of the factors that may affect the clinical prognosis of chondroblastoma patients. We also suggest future prospective controlled studies with large samples to improve the limitations encountered by Outani et al. (World J Surg Oncol. 18(1):47, 2020) due to insufficient statistical power of variables and lack of controls.

Pegylated filgrastim is comparable with filgrastim as support for commonly used chemotherapy regimens: a multicenter, randomized, crossover phase 3 study.

The purpose of this study was to compare the efficacy and safety of a single subcutaneous injection of pegylated filgrastim with daily filgrastim as a prophylaxis for neutropenia induced by commonly used chemotherapy regimens. Fifteen centers enrolled 337 chemotherapy-naive cancer patients with normal bone marrow function. All patients randomized into AOB and BOA arms received two cycles of chemotherapy. Patients received a single dose of pegylated filgrastim 100 µg/kg in cycle 1 (AOB) or cycle 2 (BOA) and daily doses of filgrastim 5 µg/kg/day in cycle 1 (BOA) or cycle 2 (AOB). Efficacy and safety parameters were recorded. The primary end point was the rate of protection against grade 4 neutropenia after chemotherapy [defined as the rate at which the absolute neutrophil count (ANC) remained >0.5×10(9)/l throughout the entire cycle]. Ninety-four percent of patients receiving pegylated filgrastim or filgrastim did not develop grade 4 neutropenia. The incidence of ANC<1.0×10(9)/l was 16.0% (50/313) after support with either pegylated filgrastim or filgrastim. The incidences of febrile neutropenia and antibiotic administration were similar in both groups. Notably, faster ANC recovery was observed with pegylated filgrastim support. The ANC nadir was also earlier with pegylated filgrastim (day 7) support than with filgrastim support (day 9), although the depth of nadir was not significantly different. A single subcutaneous injection of pegylated filgrastim 100 µg/kg provided adequate and safe neutrophil support comparable with daily subcutaneous injections of unmodified filgrastim 5 µg/kg/day in patients receiving commonly used standard-dose mild-to-moderate myelosuppressive chemotherapy regimens.

Pharmacokinetic characteristics of vincristine sulfate liposomes in patients with advanced solid tumors.

AIM: To evaluate the single- and multiple-dose pharmacokinetics of vincristine sulfate liposomes (VSLI) in patients with advanced solid tumors. METHODS: In single-dose pharmacokinetic study, 16 patients were administered VSLI (1.5, 2.0, or 2.3 mg·m(-2)) through intravenous infusion. Another 6 patients receiving vincristine sulfate (VCR, 2.0 mg) were taken as the control. In multiple-dose pharmacokinetic study, 12 patients were administered VSLI (1.5 or 1.8 mg·m(-2)) through intravenous infusion weekly for 4 consecutive weeks. The plasma concentration of VSLI was determined using the liquid chromatography-tandem mass spectrometry (LC-MS/MS) method. RESULTS: After intravenous infusion of the single dose of VSLI, the plasma concentrations were characterized by bi-exponential decline curves. No statistically significant differences were observed between the main pharmacokinetic parameters in the 3 dose groups. Compared with the patients receiving VCR, the patients treated with VSLI displayed an increase in the area under the plasma concentration vs time curve (AUC), and a decrease in plasma clearance rates. On the 4th cycle in the multiple-dose study, the plasma concentration of VCR in all subjects prior to the weekly administration was below the lower limit of quantification (LLOQ). The calculated pharmacokinetic parameters from the subjects in the multiple- and single-dose (1.5 mg·m(-2)) groups had no significant differences. Although the administration of liposomal VCR may significantly elevate the plasma concentration of VCR, VSLI-associated adverse events were similar to those associated with conventional VCR. CONCLUSION: VSLI exhibits a lower clearance and a higher AUC compared with conventional VCR. No accumulation was observed in patients exposed to VSLI for 4 consecutive weeks. VSLI was generally tolerated in the subjects. The phase II dose of VSLI may be recommended as 4 doses of 1.5 mg·m(-2) for treatment of patients with advanced solid tumors.

[Efficacy and adverse effets of nimotuzumab plus palitaxel liposome and carboplatin in the treatment for advanced non-small cell lung cancer].

OBJECTIVE: To evaluate the efficacy of nimotuzumab combined with palitaxel liposome and carboplatin (LP) regimen for treatment of advanced non-small cell lung cancer (NSCLC), and to observe the changes of tumor markers and toxicities in the treatment. METHODS Forty-one patients with advanced NSCLC were randomly divided into 2 groups: 21 patients in the observation group were treated with nimotuzumab (200 mg per week for 6 weeks), palitaxel liposome 160 mg/m2 and carboplatin (AUC = 6). 20 patients in the control group were given LP regimen. Each group completed two cycles of chemotherapy. The level of tumor markers (CEA, CYFR21-1 and NSE) and toxicities were checked at one week before and after the treatment. Thoracic CT examinations were taken before treatment and at the fourth week and eighth week after treatment. RESULTS: In the observation group, there were 2 cases of CR, 7 cases of PR, 9 cases of SD and 3 cases of PD. The objective response rate (RR) was 42. 9% in the observation group. In the control group, there were 1 case of CR, 6 cases of PR, 8 cases of SD and 5 cases of PD, with a RR of 35.0% in this group. There was no significant difference in the RR between the two groups (P = 0.751). The time to progression (TIP) was 6. 9 months in the observation group and 5. 7 months in the control group, with a significant difference (P = 0.027). The levels of NSE decreased significantly in both groups and showed a significant difference (P = 0.039). The levels of CEA and CYFRA21 in both groups were decreased after treatment, but did not show a significant difference before and after treatment, respectively. Except 3 cases had I-II skin toxicities on the faces in the observation group, there was no significant difference in toxicities between the two groups. CONCLUSION: Nimotuzmab combined with LP regimen shows a synergistic effect, can increase the efficacy and prolong TFP in advanced NSCLC patients. The toxicities are mild and tolerable.

Selective Detection of Alkaline Phosphatase Activity in Environmental Water Samples by Copper Nanoclusters Doped Lanthanide Coordination Polymer Nanocomposites as the Ratiometric Fluorescent Probe.

In this paper, a novel, accurate, sensitive and rapid ratiometric fluorescent sensor was fabricated using a copper nanoclusters@infinite coordination polymer (ICP), specifically, terbium ion-guanosine 5'-disodium (Cu NCs@Tb-GMP) nanocomposites as the ratiometric fluorescent probe, to detect alkaline phosphatase (ALP) in water. The fluorescence probe was characterized by scanning electron microscopy, transmission electron microscopy, X-ray photoelectron spectroscopy, and Fourier transform infrared spectroscopy. The experimental results showed that, compared with Tb-GMP fluorescent sensors, Cu ratiometric fluorescent sensors based on NCs encapsulated in Tb-GMP had fewer experimental errors and fewer false-positive signals and were more conducive to the sensitive and accurate detection of ALP. In addition, the developed fluorescent probe had good fluorescence intensity, selectivity, repeatability and stability. Under optimized conditions, the ratiometric fluorescent sensor detected ALP in the range of 0.002-2 U mL-1 (R2 = 0.9950) with a limit of detection of 0.002 U mL-1, and the recovery of ALP from water samples was less than 108.2%. These satisfying results proved that the ratiometric fluorescent probe has good application prospects and provides a new method for the detection of ALP in real water samples.

Characteristics and Comparisons of Morphometric Measurements and Computed Tomography Hounsfield Unit Values of C2 Laminae for Translaminar Screw Placement Between Patients With and Without Basilar Invagination.

OBJECTIVE: Patients with basilar invagination (BI) had high incidences of vertebral variations and high-riding vertebral artery (HRVA) that might restrict the use of pedicle or pars screw and increase the use of translaminar screw on axis. Here, we conducted a radiographic study to investigate the feasibility of translaminar screws and the bone quality of C2 laminae in patients with BI, which were compared with those without BI as control to provide guidelines for safe placement. METHODS: In this study, a total of 410 patients (205 consecutive patients with BI and 205 matched patients without BI) and 820 unilateral laminae of the axis were included at a 1:1 ratio. Comparisons with regard to insertion parameters (laminar length, thickness, angle, and height) for C2 translaminar screw placement and Hounsfield unit (HU) values for the assessment of the appropriate bone mineral density of C2 laminae between BI and control groups were performed. Besides, the subgroup analyses based on the Goel A and B classification of BI, HRVA, atlas occipitalization, and C2/3 assimilation were also carried out. Furthermore, the factors that might affect the insertion parameters and HU values were explored through multiple linear regression analyses. RESULTS: The BI group showed a significantly smaller laminar length, thickness, height, and HU value than the control group, whereas no significant difference was observed regarding the laminar angle. By contrast, the control group showed significantly higher rates of acceptability for unilateral and bilateral translaminar screw fixations than the BI group. Subgroup analyses showed that the classification of Goel A and B, HRVA, atlas occipitalization, and C2/3 assimilation affected the insertion parameters except the HU values. Multiple linear regression indicated that the laminar length was significantly associated with the male gender (B = 0.190, p < 0.001), diagnoses of HRVA (B = -0.109, p < 0.001), Goel A (B = -0.167, p < 0.001), and C2/3 assimilation (B = -0.079, p = 0.029); the laminar thickness was significantly associated with the male gender (B = 0.353, p < 0.001), diagnoses of HRVA (B = -0.430, p < 0.001), Goel B (B = -0.249, p = 0.026), and distance from the top of odontoid to the Chamberlain line (B = -0.025, p = 0.003); laminar HU values were significantly associated with age (B = -2.517, p < 0.001), Goel A (B = -44.205, p < 0.001), Goel B (B = -25.704, p = 0.014), and laminar thickness (B = -11.706, p = 0.001). CONCLUSION: Patients with BI had narrower and smaller laminae with lower HU values and lower unilateral and bilateral acceptability for translaminar screws than patients without BI. Preoperative 3-dimensional computed tomography (CT) and CT angiography were needed for BI patients.

[Orofacial myofunctional therapy improves facial morphology of children with obstructive sleep apnea after adenotonsillectomy].

PURPOSE: This study investigated the effectiveness of orofacial myofunctional therapy(OMT) in improving facial morphology of children with obstructive sleep apnea (OSA) after adenotonsillectomy (AT). METHODS: Ten children aged from 4-7 years with persistent oral breathing for more than 1 month after adenotonsillectomy were chosen to receive orofacial myofunctional therapy. The patients were required to take photos before and after orofacial myofunctional therapy. In order to compare the soft changes before and after OMT treatment, twelve representative mark points were selected and used for proportion and angle measurements. Graphpad Prism 8 statistical software was used for statistical analysis, to compare the differences in facial morphology of patients before and after treatment. RESULTS: Compared with before OMT, a significant difference was found in the proportion of Sn-Ls/Sn-Stms(P=0.0002), Sn-Stms/Sn-Me'(P<0.05), as well as in the angle of Gs-Sn-Pos (P<0.05), nasolabial angle(P=0.0005), mentolabial angle (P=0.0026) after OMT treatment. CONCLUSIONS: Orofacial myofunctional therapy can be considered as an effective complementary treatment for OSA patients with oral breathing after adenotonsillectomy.

Inhibitory effects of polystyrene microplastics on caudal fin regeneration in zebrafish larvae.

Microplastic pollution is pervasive in aquatic environments, but the potential effects of microplastics on aquatic organisms are still under debate. Given that tissue damage is unavoidable in fish and the available data mostly concentrate on healthy fish, there is a large chance that the ecotoxicological risk of microplastic pollution is underrated. Therefore, in this study, the effects of microplastics on the regenerative capacity of injured fish were investigated using a zebrafish caudal fin regeneration model. After fin amputation at 72 h post fertilization, the larvae were exposed to polystyrene microplastics (0.1-10 mg/L) with diameters of 50 or 500 nm. Microplastic exposure significantly inhibited fin regeneration, both morphologically and functionally. Furthermore, the signaling networks that regulate fin regeneration, as well as reactive oxygen species signaling and the immune response, both of which are essential for tissue repair and regeneration, were altered. Transcriptomic analyses of the regenerating fin confirmed that genes related to fin regeneration were transcriptionally modulated in response to microplastic exposure and that metabolic pathways were also extensively involved. In conclusion, this study demonstrated for the first time that microplastic exposure could disrupt the regenerative capacity of fish and might eventually impair their fitness in the wild.

[Experimental study of low molecular weight heparin drug delivery system for prevention of posterior capsular opacification in rabbit eyes].

OBJECTIVE: To investigate the safety and efficacy of low-molecular-weight heparin drug delivery system (LMWH DDS) for prevention of posterior capsular opacification (PCO) in rabbit eyes. METHODS: (1) To prepare the LMWH DDS by freeze-drying way with Polylactic-co-glycolic acid (PLGA) as the carrier, and evaluate its release properties in vitro. (2) Fifty New Zealand albino rabbits (50 eyes) undergoing phacoemulsification were equally divided into five groups: receiving normal saline eye drops (group A), 3 different dose (1 mg, 0.5 mg and 0.25 mg) of LMWH DDS respectively implanted into the posterior chamber (group B, C and D), and a carrier DDS implanted into the posterior chamber (group E). All the 50 eyes were examined by slit-lamp microscopy. The low-molecular-weight heparin levels in aqueous humor were measured, and the wet posterior capsules were weighed. RESULTS: The LMWH DDS prepared with a freeze-dried way has high encapsulation efficiency, and the equation of 49-day release curve fitting in vitro were were similar to zero order. The fibrin exudation in group B, C and D were lower than in Group A and E during the first postoperative day. There were 10, 2, 3, 9 and 10 eyes that developing PCO in the group A, B, C, D and E, respectively. The mean wet-weight of the posterior capsule were (114.59 +/- 14.58) mg, (24.14 +/- 6.08) mg, (39.23 +/- 17.13) mg, (99.35 +/- 29.37) mg, (115.29 +/- 19.87) mg respectively in 5 trial groups. There were stable and high concentration of low molecular weight heparin in aqueous of group B and C during the 4 weeks (> 20 mg/L), while a instable and lower concentrations in group D. The result of optical microscopy and electron microscopy examination indicated that fibroblast proliferation was quite active in groups A, D and E, but inactive in group B and C. Neither infiltration of inflammatory cells at the cornea, iris, trabecular meshwork and ciliary body nor retinal degeneration or necrosis was found in any group at 12 weeks. There was no intraocular bleeding in all the five groups in the following 12 weeks. CONCLUSIONS: The LMWH DDS prepared by freeze-drying way with PLGA as the carrier has good slow-release and biological tolerance. Implantation of LMWH DDS into the posterior chamber of experimental animals can significantly reduce postoperative fibrin exudation, and can safe and effective prevent the occurrence of PCO, and also there were some dose-effect relationship.

[Application of CAD/CAM techniques in mandible large-scale defect and reconstruction with vascularized fibular bone graft].

OBJECTIVE: To exploit computer-aided design and computer-aided manufactured (CAD/CAM) techniques and application in the reconstruction of mandible large-scale defect with vascularized fibular bone graft. METHODS: Before actually performing surgery, three-dimensional(3D) computed tomography(CT) was performed in 7 patients with mandibular large-scale defects, and 3D CT images were acquired by processing CT data. Then the CT data were transformed into a readable format and transferred to produce facsimile models by means of using rapid prototyping(RP) techniques. When individual mandibular models and enantiomorphous models were produced, evaluation and surgical simulation was performed in model, which included measuring range of mandible lesions, prefabrication of mandibular reconstructive titanium palate, precise position of titanium screws, shaping the free vascularized fibula by mandibular, etc. According to the simulations, the mandible reconstructions were finished in operation. RESULT: CAD/CAM techniques and application can distinctly display the mandibular lesions and ambient relationships, which is very useful for clinical assessment and surgical planning. Particular advantages were the unlimited trials with the imaging method, and the feeling of reality with the model method. The actual operative time was shortened, and surgery results were satisfactory with few complications. CONCLUSION: CAD/CAM techniques are very helpful for simulation of mandible large-scale defect with complicated anatomical and reconstructive problems. By preoperative simulation of procedures, surgeons can improve or refine treatment planning using this method and improve postoperative results.

[Treatment of chemotherapy-induced neutropenia pegylated recombinant human granulocyte colony-stimulating factor: a multi-center randomized controlled phase II clinical study].

OBJECTIVE: To compare the efficacy and safety of daily administration of recombinant human granulocyte colony-stimulating factor (rhG-CSF), and a single subcutaneous injection of pegylated recombinant human granulocyte colony-stimulating factor (PEG-rhG-CSF), a sustained-duration rhG-CSF, in chemotherapy-induced neutropenia. METHODS: In the present randomized, open-label, match and cross-over study, enrolled 104 patients with previously untreated non-small cell lung cancer (NSCLC), breast cancer or non-Hodgkin's lymphoma and with normal bone marrow function from 13 centers were randomly divided into 2 matched groups, AB and BA group. Each patient received two cycles of chemotherapy of identical regimen. In the study cycle, the patients received a single subcutaneous injection of PEG-rhG-CSF 100 microg/kg on day 3; and in control cycle, daily subcutaneous infection of rhG-CSF 5 microg x kg(-1) x d(-1) began on day 3 and continued for 14 days or until the absolute neutrophil count (ANC) became > or = 5.0 x 10(9)/L twice after it decreased to the nadir. Efficacy and safety parameters were monitored. RESULTS: The incidence rates of ANC < 1.5 x 10(9)/L in the 103 evaluable study cycles and 100 evaluable control cycles were 30.00% and 20.00% with the duration of 2.39 days and 2.35 days respectively. The incidence rates of grade 3 neutropenia were 7.77% and 7.00%; and that of grade 4 neutropenia were 5.80% and 4.00% respectively in the trial and control cycles. However, all the difference mentioned above did not reached statistical significance. None of the patients experienced febrile neutropenia. The ANC nadir was (7.55 +/- 5.25) x 10(9)/L and (8.42 +/- 5.57) x 10(9)/L (P = 0.257) respectively after receiving PEG-rhG-CSF and rhG-CSF. Compared with that of rhG-CSF group, the ANC profile of PEG-rhG-CSF group exhibited limited "overshoot" of neutrophils after the nadir. Subgroup analysis according to disease type yielded similar results. The safety profiles of the PEG-rhG-CSF and rhG-CSF groups were similar. Musculoskeletal pain or arthralgia occurred in 16.5% of the study cycles and 26.00% of the control cycles (P = 0.963), mostly mild or moderate. Other adverse effects such as fever, fatigue, dizziness, gastrointestinal effects and injection-site pain, were transient and easily manageable. CONCLUSION: A single subcutaneous injection of PEG-rhG-CSF 100 microg/kg provides neutrophil support and a safety profile comparable to regimen of daily subcutaneous injection of rhG-CSF 5 microg x kg(-1) x d(-1) in Chinese patients receiving a variety of myelosuppressive chemotherapy regimens.

Graphene oxide and sulfonated polyanion co-doped hydrogel films for dual-layered membranes with superior hemocompatibility and antibacterial activity.

In this study, a new kind of hemocompatible and antibacterial dual-layered polymeric membrane was fabricated by coating a top layer of graphene oxide and a sulfonated polyanion co-doped hydrogel thin film (GO-SPHF) on a bottom membrane substrate. After a two-step spin-coating of casting solutions on glass plates, dual-layered membranes were obtained by a liquid-liquid phase inversion method. The GO-SPHF composite polyethersulfone (PES) membranes (PES/GO-SPHF) showed top layers with obviously large porous structures. The chemical composition tests indicated that there were abundant hydrophilic groups enriched on the membrane surface. The examination of membrane mechanical properties indicated that the composite membranes exhibited only slightly decreased performance compared to pristine PES membranes. Moreover, to validate the potential applications of this novel dual-layered membrane in diverse fields, we tested the hemocompatibility and antibacterial activity of the membranes, respectively. Notably, the PES/GO-SPHF membranes showed highly improved in vitro hemocompatibility, such as good anti-coagulant activity, suppressed platelet adhesion and activation, low inflammation potential, and good red blood cell compatibility. Furthermore, the dual-layered membranes exhibited robust antibacterial ability after in situ loading of Ag-nanoparticles with excellent bactericidal capability to both Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus). Due to the integration of the porous membrane structure, good mechanical strength, excellent hemocompatibility, as well as robust bactericidal capability, the GO and sulfonated polyanion co-doped dual-layered membranes may open up a new protocol to greatly demonstrate the potential application of polymeric membranes for clinical hemodialysis and many other biomedical therapies.

Evaluation of toxicity and single-dose pharmacokinetics of intravenous ursolic acid liposomes in healthy adult volunteers and patients with advanced solid tumors.

OBJECTIVE: The purpose of this study was to investigate the maximum tolerated dose (MTD), dose-limiting toxicity (DLT), and pharmacokinetics of ursolic acid liposomes (UAL), as a new drug, in healthy adult volunteers and patients with advanced solid tumors. METHODS: All subjects received a single-dose of UAL (11, 22, 37, 56, 74, 98, and 130 mg/m(2)) administered as a 4-h intravenous infusion. Toxicity was assessed and plasma samples were analyzed using validated ultra-performance liquid chromatograph/tandem mass spectroscopy method. RESULTS: A total of 63 subjects including 4 patients and 35 healthy adult volunteers for toxicity study and 24 healthy adult volunteers for pharmacokinetic study were enrolled in this trial. The DLT was encountered at 74, 98, and 130 mg/m(2), and consisted of hepatotoxicity and diarrhea. Other adverse events included grade 1 nausea, grade 2 abdominal distention, grade 1 microscopic hematuria, grade 2 elevated serum sodium, grade 1 vascular stimulation, and grade 1 skin rash. The MTD was 98 mg/m(2). The single-dose pharmacokinetic parameters revealed a linear relationship between C(max), AUC(0→24 h), or AUC(0→∞) and escalated doses. CONCLUSIONS: The clinical data reported for the first time that UAL had manageable toxicities with MTD of 98 mg/m(2). The DLT were hepatotoxicity and diarrhea. Meanwhile, UAL had a linear pharmacokinetic profile. The registration number of this trial is ChiCTR-ONC-12002385.

[Application of alpha-cyanoacrylate medical adhesive in fixation of intratemporal facial nerve within chitin chamber].

OBJECTIVE: To study the feasibility of alpha-cyanoacrylate medical adhesive in fixation of intratemporal facial nerve when nerve was repaired within chitin chamber, and to investigate the nerve regeneration. METHODS: Nerve defect of 6 mm was made in left intratemporal facial nerves of 48 rabbits. All the defects were bridged with chitin chamber and were fixed by alpha-cyanoacrylate medical adhesive, surgical suture and natural union. Nerve function test and histomorphological examination were carried out at 1 month and 3 months after repair. RESULTS: It was observed that the nerve was fixed firmly to the chamber with no crack or crease by alpha-cyanoacrylate medical adhesive. The regenerated new nerve fibers were more regular and denser and the neurological function recovered much better in the group fixed by alpha-cyanoacrylate medical adhesive than in the groups those fixed by surgical suture and natural union. CONCLUSION: The medical adhesive is strong in adhesion and beneficial to nerve repair; repair of intratemporal facial nerve defect within chitin chamber fixed by alpha-cyanoacrylate medical adhesive is feasible, simple and timesaving.

[Expression of proliferating cell nuclear antigen/bcl-2 in gray gingival tissue].

OBJECTIVE: To study the expression of PCNA/bcl-2 protein in gray and normal gingival tissue, and to investigate the biological effect of procelain-fused-to-metals (PFM). METHODS: Gray gingival tissue animal model was established by PFM prosthesis and immunohistochemistry S-P method was used to detect the PCNA/bcl-2 protein expression in gingival tissue after 3 months, 6 months in PFM groups and control group. RESULTS: The expression of PCNA had significant difference between the 3th month group and the control group (P < 0.05); no significant difference was found between the 6th month group and the control group (P > 0.05). The expression of bcl-2 had no significant difference between the 6th month group and the control group (P > 0.05). CONCLUSION: There were a correlation between the expression of PCNA and inflammation of the gray gingival tissue; The expression of PCNA and bcl-2 protein does not indicate that gray gingival tissue had dysplasia change.