ObjectiveTo observe the disease-causing genes and the inheritance in sporadic retinitis pigmentosa (sRP) in Ningxia region. Methods49 sRP patients and 128 family members were recruited for this study. All the patients and family members received complete ophthalmic examinations including best corrected visual acuity, slit-lamp microscope, indirect ophthalmoscopy, fundus color photography, visual field, optic coherence tomography, full view electroretinogram. DNA was extracted from patients and family members. Using exon combined target region capture sequencing chip to screen the 230 candidate disease-causing gene mutations, polymerase chain reaction and direct sequencing were used to confirm the disease-causing mutations. Results24/49 patients (49.0%) had identified disease-causing genes, totally 16 genes were involved. There were 41 mutation sites were found, including 32 new mutations (78.0%). The disease-causing genes include USH2A, C2orf71, GNGA1, RPGR1, IFT140, TULP1, CLRN1, RPE65, ABCA4, GUCA1, EYS, CYP4V2, GPR98 and ATXN7. Based on pedigree analysis, 20 patients were autosomal recessive retinitis pigmentosa, 3 patients were autosomal dominant retinitis pigmentosa and 1 patient was X linked retinitis pigmentosa. 3/7 patients with USH2A mutations were identified as Usher syndrome. ConclusionsUSHZA is the main disease-causing of sRP patients in Ningxia region. 83.3% of sRP in this cohort are autosomal recessive retinitis pigmentosa.
Retinitis pigmentosa is a hereditary disease which is characterized by damage in retinal photoreceptor cells and retinal pigment epithelium. Its main clinical features include low vision with night blindness, progressive visual field defects, and abnormal electroretinograms. The development of gene sequencing, the diagnosis and treatment methods of retinitis pigmentosa update year by year, including gene therapy, stem cell therapy, optogenetic therapy, etc. However, there is still a big gap in these treatments from laboratory technology into effective clinical treatment drugs. Some problems which include immune response, potential mutagenesis and tumorigenesis of the inserted region, genetic toxicity, quality and stability of gene technology and stem cell technology, mass production and promotion of clinical grade drugs, and optimization of the effectiveness of drugs and surgery, etc, remain to be solved by researchers.
Retinitis pigmentosa (RP) is a group of hereditary blinding fundus diseases caused by abnormalities in photoreceptors of the retina. RP is highly heterogeneous in hereditary and cdinical phenotypes. It can be divided into simple type RP and syndrome type RP. The main inheritance patterns are autosomal dominant, autosomal recessive inheritance and X-linked inheritance. With the popularization and clinical application of gene sequencing technology, more and more disease-causing genes have been discovered, and these genes are mainly expressed in photoreceptor cells and retinal pigment epithelial cell. ln-depth understanding of RP pathogenic genes not only provides a theoretical basis for RP diagnosis and genetic counseling, but also provides guidance for RP gene therapy.
ObjectiveTo identify the causative genes of the posterior microphthalmia-retinal pigment degeneration family. MethodsA retrospective clinical study. One child (proband) and 3 family members of a family with posterior microphthalmia-retinitis pigmentosa diagnosed by clinical and genetic examination at Henan Provincial People's Hospital in July 2019 were included in the study. Medical history and family history, and draw pedigree of the patients was collected. Visual acuity, visual field, fundus color photography, optical coherence tomography and electroretinogram (ERG) were examined. The peripheral venous blood of the proband, his parents and sister, and extract the whole genome DNA was collected. Whole-exome sequencing was used to detect genetic variations, the suspected pathogenic variations were verified by Sanger sequencing, and the pathogenicity was determined by bioinformatics analysis. ResultsThe parents discovered the proband was poor vision at the age of 10 months. At the age of 3, the best corrected visual acuity of the right eye and the left eye were 0.3 and 0.4, respectively. No abnormality was found in anterior segment. Extremely high hyperopia in both eyes. The axial length was 14.47 mm and 15.78 mm, respectively. The optic disc of both eyes was relatively small and flushed, retinal folds can be observed in macular area, and no obvious pigment deposition was found. ERG examination showed that the rod system response and the maximal combined response of both eyes decreased slightly to moderately, and the single-flash cone response and the 30 Hz flicker response decreased moderately to severely. Genetic analysis revealed two novel mutations in the membrane frizzled-related protein (MFRP) gene in the proband: c.363delC/p.Thr121Thrfs*16, c.1627C＞T/p .Gln543Stop,37 in exon 4 and 13, the former was a frameshift mutation, encoding 16 amino acids and then terminated, and the latter was an nonsense mutation, truncated 37 amino acids, both which were predicted to be pathogenic and segregate with disease. The mother and sister carried c.363delC, and the father carried c.1627C＞T. ConclusionMFRP gene c.363delC/p.Thr121Thrfs*16, c.1627C＞T/p.Gln543Stop, 37 compound heterozygous mutation may be the pathogenic gene of this family.
Retinitis pigmentosa (RP) is a disease that seriously affects vision. It mainly affects rod cells and causes night blindness. At the end of the disease, due to the simultaneous involvement of cone cells, the patient’s central vision and peripheral vision loss are not effective. There is no effective treatment method. However, some studies have found that although the function of photoreceptors is lost in the pathological process of RP, the function of bipolar cells and ganglion cells and the neural connection with the visual center are preserved, which provides a condition of therapeutic application in optogenetics for optogenetics. Optogenetics controls the excitability of neurons by expressing the light-sensitive protein represented by rhodopsin ion channel protein-2 on neurons, and has shown great application prospects in reshaping the photoreceptor function of the retina. The treatment of a type of retinal degenerative disease provides an effective treatment option.
ObjectiveTo explore the light response, retinal inflammation and apoptosis of the retinal ganglion cells (RGCs) 1 year after the new type of channelrhodopsin PsCatCh2.0 was transfected into the retina of rd1 mice. MethodsTwenty-four male rd1 mice were randomly divided into rd1 experimental group and rd1 control group, 12 mice in each group. 1.5 μl of recombinant adeno-associated virus (rAAV)2/2-cytomegalovirus (CMV)-PsCatCh2.0-enhanced green fluorescent protein (EGFP) was injected into the vitreous cavity 1 mm below the corneoscleral limbus of mice in the rd1 experimental group, and the same dose of recombinant virus was injected 2 weeks later at temporal side 1 mm below the corneoscleral limbus. One year after virus injection, the light response of RGCs expressing PsCatCh2.0 was recorded by patch clamp technique; the expression of PsCatCh2.0 in the retina was evaluated by immunofluorescence staining; the transfection efficiency of recombinant virus was evaluated by the transfection efficiency of virus and the number of RGCs. Hematoxylin-eosin staining was performed to measure the inner retinal thickness. Western blotting was used to detect the protein expression of nuclear factor (NF)-κB p65 in retina; real-time quantitative polymerase chain reaction was used to detect the relative expression of tumor necrosis factor (TNF)-α, interleukin (IL)-6 and Bax mRNA. Terminal deoxynucleotidyl transferase kit was used to observe the apoptosis of retinal cells in each group of mice. ResultsOne year after the intravitreal injection of recombinant virus, PsCatCh2.0-expressing RGCs can still generate 30 pA photocurrent. The virus PsCatCh2.0-EGFP was mainly transfected into RGCs, and partly transfected into amacrine cells, almost no transfection was seen in bipolar and horizontal cells. There were no significant differences in the number of RGCs and thickness of the inner retina between the rd1 experimental group and the rd1 control group (F=14.35, 0.05; P＞0.05), while the rd1 experimental group NF-κB p65 protein expression, TNF-α and IL-6 mRNA quantification were significantly lower than those of rd1 control group (F=4.61, 5.91, 5.78; P＜0.05). The number of red fluorescent apoptotic cells in the retina of mice in the rd1 experimental group was less than that in the rd1 control group, and the Bax mRNA expression was lower than that in the rd1 control group, and the difference was statistically significant (F=7.52, P＜0.01). ConclusionOne year after intravitreal injection of recombinant virus, the PsCatCh2.0 expressing RGCs can still generate photocurrent. Long term transfection and expression of PsCatCh2.0 has no obvious cytotoxic effect on RGCs, nor it increases the inflammatory effect of the retina of rd1 mice with retinal degeneration.
ObjectiveTo observe and analyze the pathogenic genes and clinical phenotype characteristics of retinitis pigmentosa sinepigmento(RPSP). MethodsA retrospective clinical study. Two patients (proband) and five family members from two RPSP families admitted to Xiamen Eye Center of Xiamen University in December 2022 and Shenzhen Eye Hospital in July 2023 were included in the study. Two families have no blood relationship and were both Han Chinese. Detailed ocular and systemic medical history and specialized examinations were performed for all members, including color fundus photography, fundus autofluorescence (FAF), and full field electroretinogram (ff-ERG) examination. The peripheral venous blood of all members was collected, and genomic DNA was extracted. Pathogenic genes and their loci were screened using whole exome high-throughput sequencing technology. Sanger sequencing was used to verify the pathogenic genes in the two pedigrees. The pathogenicity of candidate variants was evaluated according to the American Society for American College of Medical Genetics and Genomics (ACMG) classification criteria and guidelines for genetic variants. ResultsThe two probands were male, aged 9 and 7 years, respectively. The main complaint was poor binocular vision for 6 and 3 years and poor treatment effect of amblyopia. The proband (Ⅱ2) in family 1 had a pale red color on the optic disc, with leopard-like changes in the posterior pole and thinner retinal arteries. FAF showed mottled fluorescence attenuation outside the macular vascular arch. There was no significant waveform in both bright and dark visual responses of ff-ERG. He also had 6-toed deformity of both feet, renal cysts, and a slightly overweight body. The clinical diagnosis was non-pigmentary retinitis pigmentosa. The proband of family 2 (Ⅱ1) had poor binocular vision in a dark environment and had atrophy lesions on the nasal side of the optic disc and leopard print like changes in the fundus. FAF showed uneven enhancement in the fovea. ff-ERG showed severe abnormalities in dark and light response, with significant decrease and delay in b-wave amplitude and latency. He had no other systemic abnormalities. The clinical diagnosis was binocular RPSP. There were no abnormal ocular and systemic manifestations in the two family members. Gene sequencing revealed a homozygous mutation (c.534+1G>T) of BBS2 gene, which was inherited from the mother and father respectively. Based on clinical manifestations and genetic testing results, the final diagnosis was Bardet Biedl syndrome. The genetic sequencing results confirmed a novel compound heterozygous mutation (c.950T>G: p. Leu317Arg missense mutation and c.849+1G>C splicing mutation) of BBS7 gene. His father (Ⅰ1) and mother (Ⅰ2) carried M1 heterozygous variants. Combined with the clinical manifestations and genetic testing results, the final diagnosis was Bardet-Biedl syndrome (BBS). Family 2 proband (Ⅱ1) carried the BBS7 gene C.950T>G (p.Leu317Arg) (M2) missense variation and C.849 +1G>C (M3) splice site variation. His father (Ⅰ1) and mother (Ⅰ2) carried M3 shear site variation and M2 missense variation, respectively. The two families all fit the autosomal recessive inheritance pattern, and the genotype and clinical phenotype were coseparated. According to ACMG guidelines, M1, M2 and M3 were all identified as possible pathogenic variants. ConclusionsBBS2 gene M1 homozygous variation and BBS7 gene M2, M3 complex heterozygous variation are the possible pathogenic genes in family 1 and family 2, respectively. Two families are affected by BBS and RPSP, respectively.