Dysregulation of X-ray repair cross-complementing 4 expression in the eutopic endometrium of women with endometriosis.

Recent data suggest that the DNA damage response (DDR) is altered in the eutopic endometrium (EE) of women with endometriosis and this probably ensues in response to higher DNA damage encountered by the EE in endometriosis. DDR operates in a tissue-specific manner and involves different pathways depending on the type of DNA lesions. Among these pathways, the non-homologous end joining (NHEJ) pathway plays a critical role in the repair of dsDNA breaks. The present study was undertaken to explore whether NHEJ is affected in the EE of women with endometriosis. Toward this, we focused on the X-ray repair cross-complementing 4 (XRCC4) protein, one of the core components of the NHEJ pathway. Endometrial XRCC4 protein levels in the mid-proliferative phase were found significantly (P < 0.05) downregulated in women with endometriosis, compared to control women. Investigation of a microarray-based largest dataset in the Gene Expression Omnibus database (GSE51981) revealed a similar trend at the transcript level in the EE of women with endometriosis, compared to control women. Further in vitro studies were undertaken to explore the effects of H2O2-induced oxidative stress on DNA damage, as assessed by γ-H2AX and 8-hydroxy-2'-deoxyguanosine (8-OHdG) immunolocalization, and XRCC4 protein levels in endometrial stromal (hTERT immortalized human endometrial stromal cell line (ThESCs)) and epithelial (Ishikawa) cells. A significant decrease in XRCC4 protein levels and significantly higher localization of γ-H2AX and 8-OHdG were evident in ThESCs and Ishikawa cells experiencing oxidative stress. Overall, the study demonstrates that the endometrial XRCC4 expression is dysregulated in women with endometriosis and this could be due to higher oxidative stress in endometriosis.

Endometrial DNA damage response is modulated in endometriosis.

STUDY QUESTION: Is the DNA damage response (DDR) dysregulated in the eutopic endometrium of women with endometriosis? SUMMARY ANSWER: Endometrial expression of genes involved in DDR is modulated in women with endometriosis, compared to those without the disease. WHAT IS KNOWN ALREADY: Ectopic endometriotic lesions are reported to harbour somatic mutations, thereby hinting at dysregulation of DDR and DNA repair pathways. However, it remains inconclusive whether the eutopic endometrium also manifests dysregulated DDR in endometriosis. STUDY DESIGN, SIZE, DURATION: For this case-control study conducted between 2015 and 2019, eutopic endometrial (E) samples (EE- from women with endometriosis, CE- from women without endometriosis) were collected in either mid-proliferative (EE-MP, n = 23; CE-MP, n = 17) or mid-secretory (EE-MS, n = 17; CE-MS, n = 9) phases of the menstrual cycle. This study compares: (i) DNA damage marker localization, (ii) expression of DDR genes and (iii) expression of DNA repair genes in eutopic endometrial samples from women with and without endometriosis. PARTICIPANTS/MATERIALS, SETTING, METHODS: The study included (i) 40 women (aged 31.9 ± 0.81 years) with endometriosis and (ii) 26 control women (aged 31.4 ± 1.02 years) without endometriosis. Eutopic endometrial samples from the two groups were divided into different parts for histological analysis, immunohistochemistry, RNA extraction, protein extraction and comet assays. Eighty-four genes of relevance in the DNA damage signalling pathway were evaluated for their expression in eutopic endometrial samples, using RT2 Profiler PCR arrays. Validations of the expression of two GADD (Growth Arrest DNA Damage Inducible) proteins - GADD45A and GADD45G were carried out by immunoblotting. DNA damage was assessed by immunohistochemical localization of γ-H2AFX (a phosphorylated variant of histone H2AX) and 8-OHdG (8-hydroxy-2'-deoxyguanosine). RNA sequencing data from mid-proliferative (EE-MP, n = 4; CE-MP, n = 3) and mid-secretory phase (EE-MS and CE-MS, n = 4 each) endometrial samples were scanned to compare the expression status of all the genes implicated in human DNA repair. PCNA (Proliferating Cell Nuclear Antigen) expression was determined to assess endometrial proliferation. Residual DNA damage in primary endometrial cells was checked by comet assays. Public datasets were also scanned for the expression of DDR and DNA repair genes as our RNASeq data were limited by small sample size. All the comparisons were made between phase-matched endometrial samples from women with and without endometriosis. MAIN RESULTS AND THE ROLE OF CHANCE: Endometrial expression of DDR genes and intensity of immunolocalized γ-H2AFX were significantly (P < 0.05) higher in EE, compared to CE samples. DDR proteins, especially those belonging to the GADD family, were found to be differentially abundant in EE, as compared to CE. These patterns were evident in both mid-proliferative and mid-secretory phases. Intriguingly, higher DDR was associated with increased cell proliferation in EE-MP, compared to CE-MP. Furthermore, among the differentially expressed transcripts (DETs) encoded by DNA repair genes, the majority showed up-regulation in EE-MP, compared to CE-MP. Interestingly, CE-MP and EE-MP had a comparable percentage (P > 0.05) of cells with residual DNA damage. However, unlike the mid-proliferative phase data, many DETs encoded by DNA repair genes were down-regulated in EE-MS, compared to CE-MS. An analysis of the phase-matched control and endometriosis samples included in the GSE51981 dataset available in the Gene Expression Omnibus database also revealed significant (P < 0.05) alterations in the expression of DDR and DNA repair genes in EE, compared to CE. LARGE-SCALE DATA: N/A. LIMITATIONS, REASONS FOR CAUTION: The study was conducted on a limited number of endometrial samples. Also, the study does not reveal the causes underlying dysregulated DDR in the eutopic endometrium of women with endometriosis. WIDER IMPLICATIONS OF THE FINDINGS: Alterations in the expression of DDR and DNA repair genes indirectly suggest that eutopic endometrium, as compared to its healthy counterpart, encounters DNA damage-inducing stimuli, either of higher strength or for longer duration in endometriosis. It will be worthwhile to identify the nature of such stimuli and also explore the role of higher genomic insults and dysregulated DDR/DNA repair in the origin and/or progression of endometriosis. STUDY FUNDING/COMPETING INTEREST(S): The study was supported by the Department of Biotechnology and Indian Council of Medical Research, Government of India. No conflict of interest is declared.

A Case Report of Giant Aneurysmal Bone Cyst of Distal Tibia with a Non-healing Fungating Mass and its Management in a 30-year-old Male.

Introduction: Aneurysmal bone cyst (ABC) is a benign, expansible, non-neoplastic tumor of usually long bones and is identified by blood vessels and spaces that are most often differentiated by fibrous septae. It is challenging to treat these rare, giant ABCs as they have a damaging effect on the bones and compress the nearby structures, especially in load-bearing bones of the body. Case Report: We report a case of a giant ABC in the distal tibia one-third with soft tissue component of a 30-year-old male. The patient presented to our outpatient department with complaints of pain and swelling over left ankle for 1 year. The size of the swelling was 15 cm × 10 cm × 10 cm over medial aspect of ankle with 3 discharging sinuses which present over swelling. His blood parameters were suggestive of low hemoglobin count. X-rays showed cystic lesions over medial aspect of left ankle. Computed tomography scan and magnetic resonance imaging reports were suggestive of ABC. Conclusion: Our case report is unique as it reminds us that when presented with a case of ABC, excision of fungating soft tissue with curettage followed by cementing can be a preferable and better treatment option. ABC was extensively curetted out, the formed cavity was packed with bone cement, and fixation with 3 cortico cancellous screws was carried out. At 4-month follow-up, the lesion had receded, and the patient was walking without pain and any deformity. We suggest that this method of treatment is beneficial for ABC at this site and at this age.

Case of Infected Non-Union of Distal Femur Treated with Ilizarov Fixator Assisted Knee Arthrodesis and Limb Lengthening Surgery.

Introduction: Femoral fractures in adults are around 3-6% and 0.4% of all the fractures are usually distal femoral fractures, frequently consisting of high-energy injuries which are associated with compound trauma. Conventionally, femoral-tibial fusion remains one of the last treatment choices for the recurrent septic failure. Case Report: We report a unique case where a 52-year-old male had presented with a post-operative infected non-union of distal femur and patella with discharging sinus and distal femur plate in situ. The patient presented to our outpatient department with complaints of pain and swelling over right knee with discharging sinus with fixed flexion deformity of 20° for 1-year post trauma and post-surgery. The discharging sinus was over lateral aspect of knee with purulent discharge. His blood parameters were suggestive of high erythrocyte sedimentation rate, and C-reactive protein levels and a Gram staining were suggestive of Gram-negative bacilli. X-ray showed non-union of distal femur and osteomyelitic changes and knee subluxation with distal femur plate and encirclage with K-wire for patella in situ. The patient underwents three-stage procedure of debridement with implant removal, followed by knee arthrodesis and ultimately limb lengthening surgery. Conclusion: Our case report is unique and distinctive as it shows that, when a case of infected non-union of distal femur comes with stiff and non-salvageable knee with severe arthritic changes and financial constraints, we should consider for knee arthrodesis with ilizarov ring fixator and limb lengthening surgery. Although it yielded stiff knee but with functioning limb without limb length discrepancy.

Closed Reduction of Chronically Subluxated knee with Deftix (Made in India Hexapod).

Introduction: The aim of the study was to study results of (made in India) hexapod external fixator (HEF) (deft fix)-assisted correction in a case of knee subluxation with malunited medial tibial condyle fracture. Methodology: A subject with knee subluxation was selected for application of hexapod and Ilizarov ring fixator for staged correction of subluxation using deft fix-assisted correction. Result: The study shows anatomical reduction of the subluxated knee using HEF with deft fix-assisted correction. Conclusion: The HEF does not require frame transformation and is easier to use and is known to have the ability to correct complex multiplanar deformities better and much faster than the traditional Ilizarov ring fixator which requires multiple changes in hardware while correcting complex deformities. Software-assisted hexapod correction provides more faster and accurate correction with the ability to do fine adjustments at any stage of correction.

Recurrent Giant Cell Tumor of the Distal End of the Femur.

Introduction: The aim of the study was a case report of a right-sided recurrent giant cell tumor of the distal end of the femur. Methodology: A case of a 25-year-old male patient with a history of recurrent giant cell tumor of the right distal femur who presented with chief complaints of pain over right distal femur and stiffness in the right knee for 2 years with restricted knee movement and is unable to walk. He was diagnosed with recurrent giant cell tumor of the right of the distal femur and was treated with wide excision with mega prosthesis reconstruction. Result: Wide excision with mega prosthesis reconstruction showed a good functional range of motion with early rehabilitation, stability, and mobility of joints. Conclusion: We recommend wide excision and reconstruction with mega prosthesis is an effective method compared to sandwich technique and nailing and can be successfully done in the case of recurrent giant cell tumor of the distal femur with a good outcome, functional range of motion, stability, and mobility of the joint with early rehabilitation though it is a technically demanding surgery. The knee joint could have been salvaged and the need for more extensive surgery could have been prevented, had the diagnosis of recurrent giant cell tumor was made earlier.

Panel of Autoimmune Markers for Noninvasive Diagnosis of Minimal-Mild Endometriosis.

Endometriosis, characterized by the presence of endometrial-like tissue at extrauterine sites, is a common, chronic, estrogen-dependent, inflammatory condition associated with pelvic pain, subfertility, dysmenorrhea, and dyspareunia, affecting about 10% of reproductive-age women in any population. The diagnosis of endometriosis is usually delayed on an average by 8 to 11 years leading to significant consequences in terms of disease progression. The current study was aimed to validate enzyme-linked immunosorbent assay based on the epitopes of stomatin-like protein 2, tropomodulin 3 (TMOD3), and tropomyosin 3 (TPM3) for diagnosis of minimal-mild endometriosis (revised American Fertility Society Classification (rAFS) stage I-II) and to compare the performance with the reported markers: cancer antigen (CA) 125, CA19-9, α-enolase, Serine/threonine-protein kinase (PDIK1L), and syntaxin 5. This was a cross-sectional, multicenter study conducted during the year 2012 to 2015. Women with minimal-mild endometriosis (rAFS stage I-II [n = 133]) and healthy controls (n = 104) were screened for 11 novel autoimmune markers and reported markers α-enolase, PDIK1L, syntaxin 5, CA-125, and CA19-9. The sensitivity and diagnostic accuracy of serum antibodies against all the 11 epitopes were higher than that of CA-125, CA19-9, α-enolase, PDIK1L, and syntaxin 5 for diagnosis of rAFS stage I to II endometriosis. The sensitivity of 6 biomarkers (anti-TMOD3b-autoAb, anti-TMOD3c-autoAb, anti-TMOD3d-autoAb, anti-TPM3a-autoAb, anti-TPM3c-autoAb, and anti-TPM3d-autoAb) was higher at the specificity of ≥80% for diagnosis of rAFS stage I to II endometriosis as well as ultrasound-negative endometriosis. Further, logistic regression models of this panel of biomarkers showed increase in sensitivity, specificity, and diagnostic accuracy than individual biomarkers. The panel of 6 autoimmune biomarkers could be useful in setting up of noninvasive diagnostic test for detection of minimal-mild endometriosis.