Overexpression of the PLK4 Gene as a Novel Strategy for the Treatment of Autosomal Recessive Microcephaly by Improving Centrosomal Dysfunction.

Autosomal recessive microcephaly and chorioretinopathy (MCCRP) is a neurodevelopmental disorder characterized by delayed psychomotor development, growth retardation with dwarfism, and ocular abnormalities, and its occurrence has been found to be closely related to variants of the gene encoding centrosomes. However, the association between centrosomal duplication defects and the etiology of microcephaly syndromes is poorly understood. It is well known that polo-like kinase 4 (PLK4) is a key regulator of centriole duplication, and the abnormalities of centrosomal function caused by its protein variation need to be further explored in the pathogenesis of microcephaly. In our study, we found that a patient with microcephaly and chorioretinopathy harbored compound heterozygous missense variants NM_014264.4: c.2221C > T (p.Gln741*) and NM_014264.4: c.2062 T > C (p.Tyr688His) in the PLK4 gene. Overexpression experiments of the variant PLK4 proteins then showed that the G741 variant rather than the T688H variant had lost centrosomal amplification ability, and the G741 variant but not the T688H variant induced centrosomal replication disorder, which further inhibited cell proliferation, cycle division and cytoskeleton morphology in HeLa cells. Moreover, the overexpression of the two variant proteins had inconsistent effects on the target protein PLK4 by western blot analysis, also indicating that T688H variant overexpression is not functionally equivalent to WT-PLK4 overexpression. Therefore, all data support the idea that the PLK4 mutation induces centriolar duplication disorder and reduces the efficiency of mitosis inducing cell death or cell proliferation in the etiology of microcephaly disorder.

[Genetic counseling and clinical outcome of fetus with de novo chromosomal aberrations].

OBJECTIVE: To analyze the chromosome rearrangements and clinical outcome in fetus detected at prenatal diagnosis, and provide information for genetic counseling about de novo chromosomal aberrations. METHODS: From January 2006 to December 2009, we found 12 cases of de novo chromosomal aberrations in 2 583 cases of prenatal cytogenetic analyses and reviewed the karyotypes, other experimental analyses data, fetal ultrasound findings and clinical outcomes. RESULTS: Out of the 12 de novo chromosomal aberrations, 10 had unbalanced translocations and 2 had balanced reciprocal translocations. Eight of the 10 unbalanced translocation cases were terminated therapeutically, and 2 were delivered with full term. Neonates were phenotypically normal in the 2 cases with unbalanced translocations, but 1 had language retardation when followed up. The two balanced translocation cases were delivered with full term, and the neonates were phenotypically normal and clinical examinations were normal too. CONCLUSION: Detailed cytogenetic and molecular study will be adjunctive tools for predicting the phenotype of fetus with de novo chromosomal aberrations. Fetal ultrasound examination will provide convincible demonstration to determine the outcome of pregnancy.

Macrophage migration inhibitory factor enhances neoplastic cell invasion by inducing the expression of matrix metalloproteinase 9 and interleukin-8 in nasopharyngeal carcinoma cell lines.

BACKGROUND: Nasopharyngeal carcinoma (NPC) shows highly invasive and metastatic features. This study aims to investigate macrophage migration inhibitory factor (MIF)-induced invasion of NPC cells in vitro and the effects on matrix metalloproteinases (MMPs) and interleukin-8 (IL-8), and to study the mechanism of tumor cell invasion and metastasis in the early stage of NPC. METHODS: Two nasopharyngeal carcinoma cell lines, CNE-1 and CNE-2, were adopted in this study. The NPC cell invasion and migration were evaluated by microinvasion assay. The variation of expression percentages of MMP2- or MMP9-positive cells was detected by flow cytometry in two cell lines with or without MIF treatment. Western blotting and RT-PCR were used to assay the protein and mRNA expressions of MMP2 and MMP9. The IL-8 concentration secreted by NPC cells was compared with the cells with different treatments using ELISA. RESULTS: After treating with MIF for 48 hours, the cell numbers of CNE-1 and CNE-2 which went through the 8-microm filter membrane were increased. Compared with non-MIF treated NPC cells, significant difference could be found both in CNE-1 (P = 0.005) and CNE-2 cells (P = 0.001). The percentages of MMP9-positive cells were significantly increased in both CNE-1 [from (28.5 +/- 2.5)% to (82.4 +/- 3.5)%, P = 0.001] and CNE-2 [from (32.8 +/- 3.5)% to (86.1 +/- 1.6)%, P = 0.002]. The relative intensity of MMP9 protein expression was also enhanced in both cell lines (CNE-1: from 83.1 +/- 6.0 to 242.9 +/- 22.9, P = 0.002; CNE-2: from 84.4 +/- 4.3 to 278.9 +/- 29.7, P = 0.003). Correspondingly, the increased MMP9 mRNA expression level was significantly detectable in both cell lines. The concentration of IL-8 in the supernatant of CNE-2 was higher [(1201.8 +/- 593.3) pg/ml] after treatment. It was also remarkably higher than that in the supernatant of CNE-2 without treatment (P = 0.026). However, there was no significant difference in the concentration variation of IL-8 in CNE-1 (P = 0.581), while the IL-8 mRNA level was only enhanced in CNE-2. CONCLUSIONS: MIF can induce potent invasion of NPC cell lines in vitro, and the infiltrating lymphocytes in NPC might be responsible for the invasion and metastasis of tumor cells. MIF cytokine which is secreted by these infiltrating lymphocytes might contribute to the invasion as well as metastasis of NPC in the early stages by induction of MMP9 and IL-8 in an indirect pathway.