Staining For Keratin Research Articles

A Rare Placental Site Nodule Involving the Ovary.

Placental site nodule (PSN) is a benign lesion representing a nodular aggregate of intermediate trophoblast, embedded in a hyalinized stroma, thought to arise from noninvoluted placental site remaining from a past gestation. Uterus is the most common site of PSN. Occurrence in extrauterine sites is rare, with most examples being reported in the fallopian tubes. Here we report an example of PSN in the ovary. A 35-year-old woman, gravida 4, para 1, with history of adnexal ectopic pregnancy treated with methotrexate, at 39 weeks and 1 day of a subsequent pregnancy, underwent a scheduled C-section. The surgery was successful, and a healthy female infant was delivered. Intraoperative adnexal inspection revealed a small pedunculated mass on the right ovary, which was excised and sent for pathological examination. Gross inspection showed a soft, tan-white tissue fragment measuring 2.0 × 1.0 × 0.2 cm. Microscopic examination showed epithelioid cells with hyperchromatic, mildly atypical nuclei and abundant eosinophilic cytoplasm, embedded in a hyalinized stroma, forming a nodule. A diagnosis of placental site nodule was made. Immunohistochemical stains for keratin AE1/AE3, vimentin, and inhibin were strongly positive in the epithelioid cells, and immunostain for p63 was focally positive, supporting the diagnosis. PSN of the ovary is extremely rare. To our knowledge, there has been only one reported patient in the literature so far. Extrauterine PSNs are thought to arise from ectopic pregnancies. Our patient's ovarian PSN is most likely a consequence of her previous adnexal ectopic pregnancy, which was treated medically.

Construction and Identification of a Human Colorectal Adenoma Epithelial Cell Line by SV40 Large T-Antigen Transfection.

Colorectal adenoma represents the critical step in the development of colorectal cancer. The establishment of an immortalized epithelial cell line of colorectal adenoma of human origin would provide a tool for studying the mechanism of precancerous lesions, screening the efficacy of novel drugs, and constructing in vivo disease models. Currently, there is no commercially available stable supply of epithelial cells from precancerous lesions. This study aimed to establish a natural LHPP low-expressing precancerous epithelial cell line by SV40-LT antigen gene transfection. Simian vacuolating virus 40(SV40), SV40-LT overexpressed lentivirus vector, was transfected into primary human colorectal adenomatous polyp epithelial cells. The transfected cells were screened, and the screened cells were amplified to obtain the epithelial cell line: IHCRA- CELL. The cells were identified by morphological observation, cell proliferation, Quantitative real-time PCR (qPCR), and Short Tandem Repeats (STR) experiments. Morphologically, the cells showed epithelial-like characteristics, such as polygon shape, desmosomes mitochondria, and strong positive keratin staining. There was no significant difference between the transfected cells and the primary cells. Through the STR identification experiment, no matching cell lines were found in the cell lines retrieval. We successfully established a natural LHPP low-expressing precancerous epithelial cell line by SV40-LT antigen gene transfection, which has been patented and is now preserved in the Chinese Typical Culture Preservation Center. It was verified that the transformed cells maintained the phenotype and biological characteristics of epithelial cells. This cell line can be used to study the mechanism of precancerous lesions, screen the efficacy of novel drugs, and construct in vivo disease models.

The Effects of Negative Pressure Therapy on Hair Growth of Mouse Models.

Negative pressure therapy (NPT) has been shown to facilitate wound healing and promote hair growth in a porcine model. However, there is a paucity of research on the impact of negative pressure on hair growth in murine models. Despite the ability of nude mice to develop hair follicles, the hair they produce is often flawed towing to genetically induced keratin disorders, rendering them a pertinent animal model for assessing hair regeneration. Therefore, this study aims to investigate the effects of negative pressure on hair follicle growth in a nude mouse model. To achieve this, a customized external tissue expansion device was developed to apply negative pressure to the dorsum of nude mice. The mice were subjected to several treatment courses consisting of 15 and 30 min of continuous negative pressure at 10 mmHg, which were repeated 5 and 10 times every other day until sacrifice. Dorsal skin samples were subsequently extracted from the suction and nonsuction areas. The sections were stained with various antibodies to assess the expression of SOX-9, LHX-2, Keratin-15, β-catenin, CD31, and vascular endothelial growth factor-A, and a TUNEL assay was used to analyze cell apoptosis. The results showed that the number of hair follicles and angiogenesis were significantly higher in the suction area than in the nonsuction area in all groups. Moreover, mice that received NPT for 15 min for 10 times had a higher hair follicle density than the other three groups. Immunofluorescence staining for LHX-2 and Keratin 15 further validated the results of these findings. In conclusion, this study demonstrated that negative pressure effectively promotes hair follicle growth and angiogenesis in nude mice through SOX-9- and LHX-2-mediated follicular regeneration and β-catenin-mediated hair follicle morphogenesis. Impact Statement The results of this study indicate that negative pressure therapy (NPT) is effective in promoting hair growth in nude mice, as evidenced by increased hair follicle density and angiogenesis in the treated areas. Using a custom external tissue expansion device (ETED) device, 15-min NPT treatment conducted over 10 sessions demonstrated the highest follicle density. This suggest that developing a regimen for NPT may offer to create innovative treatment approaches for hair loss, ultimately benefiting individuals suffering from hair loss disorders.

Gingival Carcinoma Cuniculatum: A Case Series of Seven Cases with Emphasis on Immunohistochemical and Histochemical Features.

Oral carcinoma cuniculatum (CC) is a rare distinct variant of oral squamous cell carcinoma (OSCC) displaying unique clinical and histopathological features. CC is a locally aggressive malignancy with lower metastatic potential and has a tendency to invade the underlying bone yielding nonspecific clinical presentation resembling fungal/bacterial infections or osteomyelitis. Seven rare cases of gingival CC are reported with special emphasis on immunohistochemistry and special stains. Clinical and demographic profile were analysed and all cases were subjected to Ki-67 and p53 immunostaining and Ayoub-Shklar histochemical staining. For comparison, age and gender matched seven cases of conventional OSCC with bony involvement were recruited. IBM SPSS statistics software version 26 (IBM Analytics, Armonk, New York, U.S.) was used to analyse the data. All cases of CC lacked p53 expression in contrast to OSCC and demonstrated intermediate to high proliferation index in stroma also. The keratin staining and cytokeratin profile in OSCC and CC were also found to be statistically different. CC is a rare well differentiated variant of OSCC with different pathways involved in the pathogenesis. Local aggressive and invasive nature could be the result of unique interaction between epithelium and stromal elements. Oral health care provider must be familiar with this uncommon variant of OSCC.

Nuclear Protein in Testis (NUT) Carcinoma: A Comprehensive Immunohistochemical Analysis of 57 Cases With Consideration of Interpretation and Pitfall Recognition.

Nuclear protein in testis (NUT) carcinoma is an aggressive carcinoma defined by NUTM1 gene rearrangement. Diagnostic challenges include morphologic overlap with poorly differentiated squamous cell carcinoma, small cell carcinoma, thoracic SMARCA4-deficient undifferentiated tumor, and other small round blue cell tumors. To comprehensively study the immunohistochemistry (IHC) features of a large cohort of NUT carcinomas. Fifty-seven NUT carcinoma cases were identified from 2012-2022, including 38 thoracic/mediastinal, 13 head and neck/sinonasal, and 6 from other sites. Pathology reports and available slides were reviewed. Comprehensive IHC studies were performed on available cases. Keratin stains showed variable positivity and were entirely negative in 15% (8 of 55) of cases. p40 was only positive in 65% (24 of 37) of cases, implying inferior sensitivity when compared to p63 (87% sensitivity, 20 of 23 cases) and other squamous cell markers. Neuroendocrine markers were focally/weakly positive in few cases; however, INSM1 was positive in 54% (7 of 13) of cases, indicating a possible diagnostic pitfall. TTF-1 was mostly negative with focal positivity in 26% (10 of 38) of cases. Occasional CD34 (15%, 3 of 20 cases) and CD99 (21%, 3 of 14 cases) positivity could also cause potential diagnostic confusion. S100, desmin, CD45, and SALL4 were rarely positive. BRG1 and INI1 were retained in all cases. Ki-67 proliferative index was high (median, 60%). PD-L1 was negative in all tested cases. This comprehensive IHC study demonstrates the immunohistochemical spectrum of NUT carcinoma. The findings can help narrow the differential diagnosis and recognize potential pitfalls.

A rare case of poorly-differentiated endometrial carcinoma in a female-to-male transgender man on testosterone therapy

Abstract Introduction/Objective Testosterone is commonly used as gender-affirming therapy for transmasculine individuals. The effects of testosterone therapy on endometrial tissue are complex, with some experiencing atrophy, and others with abnormal uterine bleeding (AUB). Endometrial carcinomas in transgender men are rarely reported in the literature. Here we present a case of poorly-differentiated endometrial carcinoma in a transgender man on testosterone. Methods/Case Report A 56 year-old female-to-male transgender man who had been on testosterone and finasteride therapy since 2015 developed vaginal discharge in 2022. No history of abnormal pap smears was noted. The patient’s pelvic ultrasound that was subsequently obtained after presenting with the vaginal discharge showed a thickened endometrium of 4 cm thickness. PET-CT revealed hypermetabolic activity corresponding to the endometrial lining of the uterus. Endometrial biopsy was notable for poorly differentiated carcinoma with retained expression of the mismatch repair proteins by immunohistochemical stains. The patient underwent hysterectomy with bilateral salpingo-oophorectomy. Grossly, there was a 9.5 cm necrotic mass occupying the entire endometrial cavity, extending >50% into myometrium thickness and extending into the lower uterine segment, without cervical involvement. Background endometrium was inactive/atrophic. Microscopic examination of the tumor showed sheets of epithelioid tumor cells with extensive necrosis, without any glandular pattern. On immunohistochemical stains, the tumor cells were positive for pankeratin and patchy staining for keratin 7 and keratin 20, without expression of PAX8, estrogen receptor, progesterone receptor or androgen receptor. Results (if a Case Study enter NA) N/A Conclusion Gender-affirming testosterone is a common long-term treatment for transgender men. There is limited understanding of the long-term effects of testosterone therapy on endometrial pathology, particularly regarding the potential for developing malignancy. Positive staining for androgen receptor has been previously reported in endometrial malignancies, and therefore, studies on the potential effects of testosterone therapy on endometrial malignancies is warranted. This case demonstrates the importance of continued gynecologic care in female-to-male transgender individuals who have not undergone previous gender affirming gynecologic surgery.

Clinicopathological and molecular features of renal cell carcinomas with haemangioblastoma-like features distinct from clear cell renal cell carcinoma.

Haemangioblastomas arise in the central nervous system. Rarely, haemangioblastomas may develop in extra-neural sites, such as the kidneys. A few reported cases of renal cell carcinomas (RCCs) with haemangioblastoma-like features have exhibited both clear cell renal cell carcinoma (CCRCC)- and haemangioblastoma-like components. The clinicopathological and molecular characteristics of RCCs with haemangioblastoma-like features were analysed, focusing on VHL alterations, in comparison with CCRCCs partially resembling haemangioblastoma. Four RCCs with haemangioblastoma-like features and five CCRCCs partially resembling haemangioblastoma were included. The RCCs with haemangioblastoma-like features were indolent and lacked adverse prognostic factors. All RCCs with haemangioblastoma-like features had a well-circumscribed appearance and a thick fibromuscular capsule, with fibromuscular bundles extending into the tumour to varying degrees in the three tumours. Each RCC with haemangioblastoma-like features exhibited CCRCC-like areas with indistinct tubular structures and foci of haemangioblastoma-like areas, in which vessels and short spindle cells overwhelmed tumour cells. Whereas haemangioblastoma-like areas in the CCRCCs partially resembling haemangioblastoma exhibited sparse vessels and spindle cells and distinct clear cells. The RCCs with haemangioblastoma-like features exhibited a unique immunohistochemical profile, with positive staining for inhibin-α, S100, carbonic-anhydrase-9, keratin7, and high molecular weight keratin and negative staining for (alpha-methylacyl-CoA racemase) AMACR. RCC with haemangioblastoma-like features did not display any VHL alterations, including VHL mutation, 3p LOH, and methylation of the VHL promoter region, and the two tumours harboured a likely oncogenic missense variant of MTOR (c.7280T>G). The histopathological, immunohistochemical, and molecular findings suggest that RCC with haemangioblastoma-like features is a distinct entity from CCRCC.

Plurihormonal PIT1-lineage pituitary neuroendocrine tumors: a clinicopathological study

Objective: To investigate the clinicopathological characteristics of plurihormonal PIT1-lineage pituitary neuroendocrine tumors. Methods: Forty-eight plurihormonal PIT1-lineage tumors were collected between January 2018 and April 2022 from the pathological database of Sanbo Brain Hospital, Capital Medical University. The related clinical and imaging data were retrieved. H&E, immunohistochemical and special stains were performed. Results: Out of the 48 plurihormonal PIT1-lineage tumors included, 13 cases were mature PIT1-lineage tumors and 35 cases were immature PIT1-lineage tumors. There were some obvious clinicopathological differences between the two groups. Clinically, the mature plurihormonal PIT1-lineage tumor mostly had endocrine symptoms due to increased hormone production, while a small number of immature PIT1-lineage tumors had endocrine symptoms accompanied by low-level increased serum pituitary hormone; patients with the immature PIT1-lineage tumors were younger than the mature PIT1-lineage tumors; the immature PIT1-lineage tumors were larger in size and more likely invasive in imaging. Histopathologically, the mature PIT1-lineage tumors were composed of large eosinophilic cells with high proportion of growth hormone expression, while the immature PIT1-lineage tumors consisted of chromophobe cells with a relatively higher expression of prolactin; the mature PIT1-lineage tumors had consistently diffuse cytoplasmic positive staining for keratin, while the immature PIT1-lineage tumors had various expression for keratin; the immature PIT1-lineage tumors showed more mitotic figures and higher Ki-67 proliferation index; in addition, 25.0% (12/48) of PIT1-positive plurihormonal tumors showed abnormal positive staining for gonadotropin hormones. There was no significant difference in the progression-free survival between the two groups (P=0.648) by Kaplan-Meier analysis. Conclusions: Plurihormonal PIT1-lineage tumor belongs to a rare type of PIT1-lineage pituitary neuroendocrine tumors, most of which are of immature lineage. Clinically increased symptoms owing to pituitary hormone secretion, histopathologically increased number of eosinophilic tumor cells with high proportion of growth hormone expression, diffusely cytoplasmic keratin staining and low proliferative activity can help differentiate the mature plurihormonal PIT1-lineage tumors from the immature PIT1-lineage tumors. The immature PIT1-lineage tumors have more complicated clinicopathological characteristics.

"Ayoub-Shklar" as a Special Stain for Keratin: A Histopathological Study.

Special stains in histopathological studies are used to identify the structures with different dyes apart from the routine stain hematoxylin and Eosin (H and E). The component which is present in the special stains will have a specific bond and affinity for particular tissue components in the histological specimen. Structures like keratin takes up an eosinophilic stain in routine (H and E) staining. Most of the potentially malignant disorders and carcinomas arise due to the keratinization defect, which makes keratin an important diagnostic tool. There are different stains such as Ayoub-Shklar (A-S), Dane-Herman (D-H), and rapid Papanicolaou (PAP) which is used to identify keratin. In A-S stain, keratins can be stained in magenta-red and orange colors. we compared A-S special stain and routine stain in terms of staining intensity or quality, the pattern of staining, and specificity for staining keratin. Thirty cases from the department archives that included 10 well-differentiated squamous cell carcinoma, 10 verrucous carcinoma, and 10 epithelial dysplasia were taken and each case was stained with both A-S and H and E stain. A-S showed an almost equal distribution of uniform and patchy staining patterns, but H and E showed more patchy staining patterns in the three groups. H and E stain shows good staining quality than A-S. A-S shows almost 90% of satisfactory staining specificity. Special stain like A-S stain can be used to stain keratin in different color, but H and E always remain gold standard stain.

Multilineage Pituitary Neuroendocrine Tumors (PitNETs) Expressing PIT1 and SF1.

PitNETs are usually restricted in their cytodifferentiation to only one of 3 lineages dictated by expression of the pituitary transcription factors (TFs) PIT1, TPIT, or SF1. Tumors that show lineage infidelity and express multiple TFs are rare. We searched the pathology files of 4 institutions for PitNETs with coexpression of PIT1 and SF1. We identified 38 tumors in 21 women and 17 men, average age 53 (range 21-79) years. They represented 1.3 to 2.5% of PitNETs at each center. Acromegaly was the presentation in 26 patients; 2 had central hyperthyroidism associated with growth hormone (GH) excess and one had significantly elevated prolactin (PRL). The remainder had mass lesions with visual deficits, hypopituitarism, and/or headaches. Tumor size ranged from 0.9 to 5cm; all 7 lesions smaller than 1cm were associated with acromegaly. Larger lesions frequently invaded the cavernous sinuses. Four cases represented a second attempt at surgical resection. PIT1 was usually diffusely positive but 5 cases had variable (patchy or focal) staining. SF1 reactivity was variable in intensity but diffuse in all but 2 cases. GATA3 data, available in 14 cases, identified diffuse positivity in 5 and focal staining in 1. GH was expressed in all but 5 tumors, PRL and thyrotropin (TSH) were expressed in 14 and 13, respectively, follicle-stimulating hormone (FSH) in 11 of 18, and luteinizing hormone (LH) in 4 of 17. Keratin staining patterns were diffuse perinuclear/membranous in 27, variable perinuclear in 4, and negative in 3; scattered fibrous bodies were seen in 5 and diffuse fibrous bodies in 1. Ki67 labeling index ranged from < 1 to 7.9%. In 3 cases, these tumors represented one of multiple synchronous PitNETs; a separate corticotroph tumor was seen in 2 patients and one patient had 2 additional discrete lesions, a sparsely granulated lactotroph, and a pure gonadotroph tumor comprising a triple tumor. PitNETs expressing PIT1 and SF1 represent multilineage PitNETs. These rare tumors have variable clinical and morphological features, most often presenting as large tumors with GH excess and occasionally as one of multiple synchronous PitNETs of distinct lineages.

Thoracic SMARCA4-deficient undifferentiated tumor: A clinicopathologic study.

e14642 Background: Thoracic SMARCA4-deficient undifferentiated tumor is a high-grade malignant neoplasm that frequently shows undifferentiated morphology and deficiency of SMARCA4. There is limited data in literature for this entity. The purpose of this report is to describe clinicopathologic features of this rare tumor in a series of 5 patients, diagnosed and treated at our institution. Methods: After approval from institutional review board, pathology department archive was searched for the patients with diagnosis of Thoracic SMARCA4-deficient undifferentiated tumor from January 2020 to December 2022. Data such as patient demographics, pathologic features including immunohistochemical stains, treatment and clinical outcome were collected. Results: Among 5 patients, 3 were male and 2 were female. Median age was 62 years (range 49 to 77 years). In three patients the primary site of the tumor was lung. One patient had tumor of the gastroesophageal junction while one patient had epicenter of the tumor in the paraspinous muscle. Diagnosis was made on biopsy specimen in all cases. All tumor showed undifferentiated histologic appearance. Rare cells with keratin positivity were observed in 2 specimens while 3 were completely negative for keratin stains. Focal TTF1 positivity was observed in one specimen while 4 were negative. Among neuroendocrine markers performed on variable number of cases, synaptophysin was positive in 4/4 cases while chromogranin was negative in 2/2 cases and INSM1 was negative in 1/1 case. PD-L1 (Dako 22C3) was performed on two specimens and showed less than 1% tumor cells to be positive. Next generation sequencing was performed on one specimen and showed pathogenic variants in TP53, CDKN2A, and APC genes and likely pathogenic variants in ATR, SMARCA4, and BCOR genes. Two patients received carbotaxol and radiation, one patient received FOLFOX, nivolumab and radiation, 1 patient pursued hospice care and 1 patient lost follow up. Two patients died 2 and 7 months after diagnosis. One patient is alive with metastatic disease 4 months after initial diagnosis. Conclusions: SMARCA4-deficient undifferentiated tumor is an aggressive malignancy that shows undifferentiated histology with synaptophysin positivity. From a diagnosis standpoint, Pathologists need to keep this entity in mind while working up tumors showing poorly differentiated to undifferentiated histology that are negative for most immunohistochemical markers. From clinical standpoint there is need to develop targeted therapy for this aggressive tumor.

Prevalence of Thyroid Transcription Factor-1 (TTF-1)-Negative Small Cell Carcinoma and Napsin A Positivity in Small Cell Carcinoma in a Cross-Sectional Study of Lung Core Biopsies.

Background The prevalence of thyroid transcription factor-1 (TTF-1) and napsin A expression are poorly characterized in lung core biopsies of small cell carcinoma. Locally, the TTF-1 clone is 8G7G3/1 (Agilent/Dako), and the napsin A clone is IP64 (Leica Biosystems). Methods All in-house lung core biopsy reports for cases accessioned at a regional laboratory from January 2011 to December 2020 were retrieved and analyzed using a validated hierarchical free-text string matching algorithm (HFTSMA) to establish the diagnosis. TTF-1 and napsin A were manually coded with the assistance of a logical text parsing tool. All TTF-1-negative small cell lung carcinoma (SCLC) cases had a full report review by pathologists. Results The cohort had 5,867 lung core biopsies, and 232 cases were confirmed as small cell carcinoma on pathologist review. TTF-1 immunostain results were available in 173 SCLC cases, and 16 cases of TTF-1-negative SCLC were confirmed on full report review. These 16 cases had at least one positive neuroendocrine (NE) marker and positive keratin staining; cases with mixed histology or positiveCK5/6 staining were excluded. Ki-67 was done in 10/16 cases; the average Ki-67 was 75%. Napsin Awas negative in 50/51 small cell carcinomas, and 0/3 TTF-1-negative SCLC had napsin A positivity. Conclusions Standardized immunostain reporting would simplify such analyses. Based on thecohort, approximately 9% (16/173) of SCLCis TTF-1 negative. Napsin A positivity in suspected small cell carcinoma should prompt consideration of an alternate diagnosis or explanation.

Unilocular thymic cyst with features of intra-thymic bronchogenic cyst: A clinicopathological correlation of 4 cases.

Four cases of primary unilocular thymic with features of bronchogenic cyst are presented. The patients are three men and a woman between the ages of 43 and 57 years. All the patients presented with symptoms of cough, chest pain, and dyspnea. Diagnostic imaging revealed the presence of an anterior mediastinal cystic lesion. Initial mediastinoscopic biopsies were non-diagnostic. Surgical resection of the anterior mediastinal cystic lesion was performed in all patients. Macroscopically, all the lesions were described as large cystic lesions measuring from 2 to 4cm in largest diameter and containing yellowish fluid. The surface of the cysts was described as corrugated and of brownish color. The walls of the cysts were not thickened and no other elements were described. Histologically, the cystic lesions were characterized by single cystic structures lined by ciliated epithelium. No atypia or mitotic activity was present in any of the cases. Immunohistochemical stains show positive staining of the respiratory epithelium with keratin and negative staining for GATA3, PAX-8, TTF-1, and Napsin. All the patients have recovered well after surgical resection.

S2919 Bile Duct Injury Due to Ketamine Use

Introduction: Ketamine is used routinely in the hospital for anesthesia, but is also used recreationally for its hallucinogenic and dissociative side effects. Chronic recreational use has been linked to bile duct damage and ulcerative cystitis. We report a case of ketamine induced sclerosing cholangiopathy. Case Description/Methods: A 42-year-old Cantonese woman with past medical history of bilateral hydronephrosis with stents leading to chronic kidney disease presented with abdominal pain associated with abnormal transaminases. Alkaline phosphatase was high at 1,017 IU/L with gamma glutamyl transferase of 2,310 IU/L with normal bilirubin. Abdominal imaging showed diffuse dilatation of the common bile duct. Viral serologies and anti-mitochondrial antibody were negative. She reported chronically elevated liver tests of unknown etiology. Social history was notable for prior alcohol abuse. She denied family history of liver diseases. Her pain resolved and plan was for outpatient follow-up, but she re-presented with sepsis with a rise in bilirubin to 2.9 mg/dL (Table 1). Given prior abnormal imaging, a magnetic resonance cholangiopancreatography was obtained, which showed increased intra- and extrahepatic bile duct dilation with irregular appearance of the central intrahepatic ducts with possible stricture. Differential diagnoses were recurrent pyogenic cholangitis, sclerosing cholangitis, and IgG4 disease. Her IgG level was high at 2,779 mg/dL with IgG4 elevated at 213 mg/dL. Liver biopsy showed chronic cholestatic liver injury consistent with sclerosing cholangitis (Image 1). IgG4 stain was negative. Upon further investigation, patient admitted to at least a decade of daily ketamine use, which was likely the culprit of both her kidney and liver diseases. Discussion: Due to her age, race, gender and rarity of ketamine induced cholangiopathy, she was not directly asked about ketamine use. Since the early 2000s, ketamine has emerged as the illicit drug of choice in Hong Kong and has seen increased use throughout Asia. The biliary damage from ketamine has been studied by a Chinese group that found 62% of the 257 chronic ketamine users had biliary tract anomalies. The greater the alkaline phosphatase, the higher the likelihood of finding biliary tract anomalies on imaging. Ketamine cessation has resulted in normalization of liver tests and imaging, however worsening cholangiopathy has been seen despite abstinence. In patients who present with unexplained cholangiopathy, chronic ketamine use should be considered.Figure 1.: Liver biopsy: A mild lymphoplasmacytic inflammation is noted in some triads with focal and mild interface activity. The bile ducts are injured (irregular contour, unevenly distributed nuclei, intracytoplasmic vacuoles) with focal periportal cholestasis. Immunohistochemical stain for keratin 7 confirms predominantly periportal cholestatic hepatocytes. Keratin 19 shows widespread loss of canals of Hering and focal bile duct loss. Rhodanine stain confirms periportal hepatocellular copper accumulation compatible with a chronic cholestatic process. Table 1. - Patient's hepatic function panels during initial encounter, subsequent encounter, and at least 6 months after subsequent encounter Liver Tests Reference Range & Units Initial Hospitalization Subsequent Hospitalization Most Recent Liver Tests Total bilirubin 0.2 – 1.2 mg/dL 0.8 2.9 4.5 Direct bilirubin 0 – 0.5 mg/dL 0.7 1.8 3.3 Aspartate aminotransferase 5 – 34 IU/L 75 104 88 Alanine transaminase 0 – 37 IU/L 74 129 91 Alkaline phosphatase 40 – 150 IU/L 1017 1106 845 Gamma glutamyl transferase 12 – 43 IU/L 2310 2516 1897

Spindle cell thymoma (WHO type A) - Choriocarcinoma: An unusual association: A clinicopathological and immunohistochemical study of 3 cases.

Three cases of an unusual association between spindle cell thymoma (WHO type A) and choriocarcinoma are presented. The patients are three men between the ages of 58 and 68 years. Clinically, all the patients presented with non-specific symptoms of cough, dyspnea, and chest pain. Clinical history and physical examination did not reveal the presence of any prior malignancy. Diagnostic imaging showed in the three patients the presence of a large anterior mediastinal mass. A core needle biopsy was obtained in the three patients. In two patients the biopsy showed the classic histology of a spindle cell thymoma while in one patient the biopsy showed the association of two tumors - spindle cell thymoma and choriocarcinoma. Surgical resection via thoracotomy was performed in the three patients. The mediastinal tumors measured between 9 and 17cm in greatest diameter and were described as solid and lobulated with areas of hemorrhage. Histologically, all the tumors showed similar histological features of spindle cell thymoma (WHO type A) associated with a high-grade neoplasm composed of round and multinucleated giant cells compatible with choriocarcinoma. Immunohistochemical stains showed positive staining for keratin 5/6, and p40 in the spindle cell component, while the choriocarcinomatous component showed positive staining for human chorionic gonadotropin and human placental lactogen. The cases herein presented highlight not only the unusual association of spindle cell thymoma and choriocarcinoma but also raises some issues regarding the histogenesis of germ cell tumors, in this case choriocarcinoma.

Epiprofin Transcriptional Activation Promotes Ameloblast Induction From Mouse Induced Pluripotent Stem Cells via the BMP-Smad Signaling Axis.

The transcriptional regulation of induced pluripotent stem cells (iPSCs) holds promise for their directed differentiation into ameloblasts, which are usually lost after tooth eruption. Ameloblast differentiation is regulated by multiple signaling molecules, including bone morphogenetic proteins (BMPs). Epiprofin (Epfn), a transcription factor, is expressed in the dental epithelium, and epithelial Epfn overexpression results in ectopic ameloblast differentiation and enamel formation in mouse incisor, a striking phenotype resembling that of mice with deletion of follistatin (a BMP inhibitor). However, it remains unknown whether and how Epfn transcriptional activation promotes ameloblast induction from mouse iPSCs. Here, we generated doxycycline-inducible Epfn-expressing mouse iPSCs (Epfn-iPSCs). Ameloblasts, which are characterized by positive staining for keratin 14 and amelogenin and alizarin red S staining, were successfully derived from Epfn-iPSCs based on a stage-specific induction protocol, which involved the induction of the surface ectoderm, dental epithelial cells, and ameloblasts at stages 1, 2, and 3, respectively. Epfn activation by doxycycline at stages 2 and/or 3 decreased cell proliferation and promoted ameloblast differentiation, along with the upregulation of p-Smad1/5/8, a key regulator of the BMP-Smad signaling pathway. Gene analysis of the BMP-Smad signaling pathway-associated molecules revealed that Epfn activation decreased follistatin expression at stage 2, but increased BMP2/4/7 expression at stage 3. Perturbations in the ameloblast differentiation process were observed when the BMP-Smad signaling pathway was inhibited by a BMP receptor inhibitor (LDN-193189). Simultaneous LDN-193189 treatment and Epfn activation largely reversed the perturbations in ameloblast induction, with partial recovery of p-Smad1/5/8 expression, suggesting that Epfn activation promotes ameloblast induction from mouse iPSCs partially by upregulating BMP-Smad activity. These results reveal the potential regulatory networks between Epfn and the BMP-Smad pathway and suggest that Epfn is a promising target for inducing the differentiation of ameloblasts, which can be used in enamel and tooth regeneration.

Lingual papillary system of the Egyptian mouse-tailed bat (Rhinopoma hardwickii): New insights to its feeding strategies using scanning electron microscope, and immunohistochemical analysis of keratin.

The current investigation described the ultrastructural and histological features of the lingual papillary system in 14 adult Rhinopoma hardwickii using scanning electron and light microscopy, and immunohistochemical staining of keratin. Three types of lingual papillae were recognized; one filiform mechanical and two gustatory (fungiform and circumvallate) of three directions: posterior, posteromedian, and median. Five filiform papillae subtypes were observed in four regions. The lingual tip had numerous giant filiform papillae. The lingual root was divided into two regions, an anterior papillary and a posterior non-papillary region that had only three circumvallate papillae in a triangular pattern. Histologically, the papillae exhibited detached thick keratin that spread over the dorsal epithelial surface. The dorsal surface of the root had stratified keratinized circumvallate papillae. Concerning the pan-cytokeratin (CK) antibody staining, moderate CK expression was observed in the superficial corneal layer of the apical lingual mucosa. While the superficial corneal cell layer in both body and root exhibited increasing of CK antibody. Morphometrically, there was a significantly increased, region-specific number of triangular, pointed filiform papillae, which were relatively increased in length and width. However, the round conical blunt filiform papillae and fungiform papillae were sparse. We concluded that our results showed distinguished adaptations in the lingual structure and its different papillae established on the nutritional and energetic demand of this bat species. This lingual adaptation is summarized in number, position, taste buds, direction, ultrastructural features, and histological that concerned with its ability in eating during the night flight. RESEARCH HIGHLIGHTS: Three types of lingual papillae; one mechanical (filiform) and two gustatory (fungiform and circumvallate) in which there were three circumvallate papillae on the posterior part of the lingual root in a triangular arrangement with an irregular surface, but the filiform papillae had five subtypes arranged in four regions. The lingual root was divided into two regions, an anterior papillary region and a posterior non-papillary region that only had three circumvallate papillae. Concerning staining with a pan-cytokeratin (CK) antibody, moderate CK expression was observed in the superficial corneal layer of the lingual mucosa of the tongue apex while the superficial corneal cell layer of the body and root exhibited increased expression of the CK antibody compared to the lingual apex.

Epithelial lip mucosa transplant re‐populates corneal anterior surface. An ex vivo study

AbstractPurposeTo establish a new ex vivo model and study the labial mucosal epithelium grafting as a potential approach for cornea re‐epithelization in patients with bilateral limbal stem cell deficiency.MethodsFour human donor corneal buttons were collected from the Eye tissue bank in corneal cold storage solution and left at 4°C for 32–52 days. Labial oral mucosa strips were dissected from four patients during fornix reconstruction after they signed an informed consent. The labial mucosa epithelial graft was prepared by dissecting substantia propria. The resulted graft was sutured near the limbus of the corneal button with running sutures (construct). Each construct was cultured under the standard conditions with the concave side upwards. After 21 days of culture, constructs were removed, washed and fixed. First half of the constructs were stained with hematoxylin and eosin (HE). 5 um crosscut cryo‐sections of the second half were stained with anti‐ keratin 13, 19, and p63, and paired secondary fluorescent antibodies. Nuclei were counterstained with Hoechst. Sections were accessed and photographed.ResultsAfter the culture, all constructs were integral with the attached epithelium graft and non‐loosened sutures. The graft became slightly flattened and brighter. HE staining demonstrated absence of the native cells in donor corneas. The labial mucosa grafts were located on the surface of the Bowman's membrane (BM) at the cornea periphery. Epithelial outgrowths were found to be transit from the labial mucosa graft to the BM. Cellular outgrows of the constructs had positive staining for keratin 13 and 19. Few cells were p63‐positive in nuclei.ConclusionsIn the study, a new ex vivo model composed of processed human donor cornea and live human donor labial mucosal epithelium graft was successfully established. The study confirmed the consistency of the new approach for corneal re‐epithelization by transplantation of an oral mucosal epithelial graft.

Primary Warthin’s-like adenocarcinoma of the lung: A clinicopathological, immunohistochemical, and molecular analysis of three cases

Three cases of primary pulmonary adenocarcinoma with prominent lymphoid stroma and papillary features mimicking Warthin’s tumor are presented. The patients are two women and one man ages 52, 62, and 74 years respectively. Clinically, the patients presented with non-specific symptoms of cough, dyspnea, and chest pain. Imaging showed the presence of an intrapulmonary mass in the right lower, right upper and left lower lobe. All patients underwent lobectomy. Histologically, the tumors were characterized by the presence of and oncocytic papillary growth pattern embedded in a lymphoid rich background. Immunohistochemical stains for TTF-1, keratin 7, and beta-catenin were positive in the epithelial component, while CD20 showed strong positive staining in the lymphoid component. In addition, CD4 and CD8 also showed positive staining in a ratio of 3–4:1, EBER was negative. Kras mutations with wild type EGFR were identified in one case. Clinical follow-up ranging from 8 to 24 months was obtained showing that all patients are alive without recurrence. The cases herein presented represent an unusual histological variant of primary lung adenocarcinoma, which closely mimics Warthin’s tumor.

Mechanisms of the mechanically activated ion channel Piezo1 protein in mediating osteogenic differentiation of periodontal ligament stem cells via the Notch signaling pathway

To explore the mechanism of Piezo1 protein in mediating the osteogenic differentiation of human periodontal ligament stem cells (hPDLSCs) via the Notch signaling pathway. In this study, young permanent teeth extracted from impacted teeth of 8-14-year- old children from January 1, 2016 to January 1, 2018 in the Department of Orthodontic, Beijing Children's Hospital were selected as cell sources. hPDLSCs were extracted by enzymatic digestion. Immunohistochemical staining was used to detect the expression of keratin and vimentin, and flow cytometry was used to identify the markers (CD146 and STRO-1) of hPDLSCs. The construction and screening of Piezo1 siRNA gene interference vector and Piezo1 gene overexpression plasmid were completed. Flexcell 4000T mechanical distraction stress instrument was used to construct hPDLSC cell model in vitro. According to the preliminary results, the experiment was divided into five groups: siRNA interference group, overexpression group, blank control group, stretch stress group, and negative control group. Real time quantitative polymerase chain reaction (RT-qPCR) was used to detect the expression of Piezo1, Notch1, alkaline phosphatase (ALP), Runt-related transcription factor 2 (Runx2), osteocalcin (OCN), and bone sialoprotein (BSP). Western blot was used to detect the expression of ALP and Runx2. Fluo-3 AM probe was used to detect intracellular calcium content. Vimentin staining of hPDLSCs was positive, and keratin staining was negative. Flow cytometry was used to detect the expression of STRO-1 and CD146, markers of hPDLSC. Empty viral vectors, siRNA-Piezo1 interference sequence, and Piezo1 overexpression vector sequence could be transfected into hPDLSC by lentivirus, and the transfection efficiency was high (approximately 90%). The reverse transcription-polymerase chain reaction (RT-PCR) results showed that there were significant differences in Piezo1 gene levels among the siRNA interference group, overexpression group, blank control group, stretch stress group, and negative control group (F=9.573, P<0.05). The level of Piezo1 in the overexpression group was significantly higher than that in the siRNA interference group (q=3.893, P<0.05). The level of Piezo1 in the stretch stress group was significantly higher than that in the blank control group (q=2.006, P<0.05). The expression of Notch1 and osteogenic genes ALP, Runx2, OCN, and BSP had the same trend. Western blot results showed that there were significant differences in the expression of ALP in the siRNA interference group, overexpression group, blank control group, stretch stress group, and negative control group (F=11.207, P<0.001). The expression level of ALP in the overexpression group was significantly higher than that in the siRNA interference group (q=2.991, P<0.05). The expression of ALP in the stretch stress group was significantly higher than that in the blank control group (q=3.007, P<0.05). The expression of Runx2 protein showed the same trend. The intracellular calcium fluorescence intensity of the overexpression group was significantly higher than that of the siRNA interference group, and the intracellular calcium fluorescence intensity of the stretch stress group was significantly higher than that of the siRNA interference group. Mechanical stretch stress can promote the expression of Piezo1 protein. Ca2+ is the second messenger, activates the Notch1 signaling pathway and the expression of ALP, Runx2, OCN, and BSP; and promotes the osteogenic differentiation of hPDLSC. The siRNA-Piezo1 interfering plasmid can block this process. On the contrary, the overexpression plasmid of Piezo1 can promote the osteogenic differentiation of PDLSCs.