Interferons in enteroviral heart disease: modulation of cytokine expression and antiviral activity

Interferon (IFN)- has a more than 120-fold higher antiviral activity than the closely related IFN- in human myocardial fibroblasts infected with the cardiotropic enterovirus coxsackievirus B3 (CVB3). CVB3 replication induces interleukin (IL)-6 and IL-8 expression in myocardial fibroblasts, and suppresses the expression of monocyte chemoattractant protein-1 (MCP-1). We investigated whether the higher antiviral activity of IFN- compared to IFN- was a result of a suppression of IL-8 expression by IFN- since previous studies had indicated that IL-8 stimulates enterovirus replication. Human myocardial fibroblasts were treated with either IFN-, IFN- or IFN- (0, 10, 100, or 1,000 IU/ml) and the concentrations of IL-6, IL-8 and MCP-1 were measured in culture supernatants by immunoassays. Both IFN- and IFN- reduced IL-6 and IL-8 expression significantly. In addition, neutralization of IL-8 in culture supernatants of myocardial fibroblasts using a monoclonal antibody demonstrated a significant reduction of CVB3 titers. Antiproliferative effects of all three IFNs were very low (<30% with 1,000 IU/ml), indicating that the suppression IL-6 and IL-8 was not related to cytotoxicity. MCP-1 expression was increased only by high concentrations of IFN- (1,000 IU/ml). By contrast, IFN- had no significant effect on IL-6, IL-8 and MCP-1 expression. In conclusion, suppression of IL-8 expression is an "immuno-modulating" feature of IFN- in human myocardial fibroblasts, which is similar to the activity of IFN-. This feature of IFN- contributes to its high antiviral activity against CVB3 and may be useful in the treatment of enteroviral heart disease.     

Glucocorticoids modulate TGF-beta production

TGF- is thought to play a central role in pulmonary fibrosis inducing fibroblast differentiation and extracellular matrix synthesis. In human lung fibroblasts, it is still unclear how various TGB- isoforms affect TGF- production and whether glucocorticoids, commonly used agents to treat fibrotic lung disease, modulate these processes. To this end, human fetal lung fibroblasts (HFL-1) were cultured with various concentrations of glucocorticoids (budesonide, dexamethasone or hydrocortisone) with and without TFG-1, -2, and -3. TGF- mRNA was assessed by real time RT-PCR. Smad 2, 3, and 4 and AP-1 complex (c-fos and c-Jun) cellular localization were evaluated by immunostaining. TGF-2 and -3 stimulated TGF-1 production significantly (p < 0.01 relative to control). TGF-1 stimulated TGF-2 production (p < 0.01 relative to control). TGF-3 was undetectable. Glucocorticoids significantly inhibited TGF-1 and -2 production and reduced expression of the upregulated TGF-1 and -2 mRNA induced by exogenous TGF-1, -2 or -3 (p < 0.01 for each) but had no effect on Smads. Although c-jun-related nuclear staining was not intensified in TGF--stimulated cells, it was reduced by glucocorticoids. Thus, TGF- isoforms may stimulate production of various TGF- isoforms in the lung. Glucocorticoids then may block TGF- production by modulating mRNA levels and c-Jun.     

Augmentation of antigen-specific cytokine responses in the early phase of vaccination with a live-attenuated simian/human immunodeficiency chimeric virus expressing IFN-γ

Summary. A nef-deleted SHIV-NM-3rN (SHIV-NI) was previously shown to be nonpathogenic and to induce protective immunity. In the present study, a SHIV-NI expressing human interferon- (SHIV-IFN-) was constructed and the effect of co-expression of IFN- on virus replication and immunopotentiation was investigated in macaques that were vaccinated with both viruses, by comparing cytokine responses during the first 4 weeks after vaccination. Peripheral blood mononuclear cells (PBMC) isolated from vaccinated macaques were stimulated with inactivated viral particles for 24h, and the production of IL-2, IL-4, IL-6, IL-10, IL-12, TNF- and IFN- was determined by ELISA and flow cytometry. All of the vaccinated macaques showed increases in cytokine production. However, the production of IFN- (Th1-type cytokine) was more rapidly induced by SHIV-IFN- vaccination, and IFN--producing cells appeared to be still increasing at 4 weeks after vaccination, although the difference of virus replication during the time was not significant in contrast to in vitro replication in cultured PBMC. These results suggest that co-expression of IFN- with SHIV can modulate the antiviral immune responses into the Th1 type response, which would probably provide more protective immunity.     

On the terms “reticulosis” and “reticulum cell sarcoma” with regard to the modern concept of the monocyte macrophage system

Summary The terms reticulosis and reticulum cell sarcoma (= malignant lymphoma, histiocytic type) are discussed regarding the modern concept of the monocyte macrophage system which today has replaced the ancient theory of the reticuloendothelial system. The monocyte macrophage system is not independent, but closely related to the myeloid system. Thus, a third blood forming system as was believed in the case of RES does not exist. Phagocytic reticulum cells of the various hematopoietic organs are highly activated monocyte-derived macrophages. All those conditions formerly termed reticuloses have been found to belong either to the myeloid or to the lymphatic system. Considering the reticulum cell sarcomas or malignant histiocytic lymphomas, most of them seem to be of lymphatic rather than of macrophage origin, representing highgrade malignant lymphomas, possibly immunoblastic sarcomas. No relationship between these tumours and the monocyte macrophage system has been established, so far. Therefore, the terms reticulosis and reticulum cell sarcoma should be no longer used in order to avoid confusion, in order to stimulate sufficient diagnostic efforts which will really clarify such cases, and in order to give full credit to modern results of hematopathology.     

Ribonuclease activity of the “myofibrillary” fraction of normal and autolyzed muscle tissue

The separation in a sucrose gradient of the myofibrillary fraction of normal and autolyzed muscle tissue gave 4 components. During post-mortem destruction of the tissue there was observed a slight decrease of the myofibrillary fraction yield and also certain changes in the distribution of protein between different components. Under the selected conditions RNase activity was found in all 4 components. During the course of autolysis enzymatic activity increased in the whole myofibrillary fraction, as well as in the lysosomal-mitochondrial components of myofibrils.Research Laboratory, Ministry of Health of the USSR, Moscow. Translated from Buylleten' Éksperimental'noi Biologii i Meditsiny, Vol. 85, No. 5, pp. 533–537, May, 1978.     

Neuron Activity in the Prefrontal Cortex of the Brain in Rats with Different Typological Characteristics in Conditions of Emotional Stimulation

Male Wistar rats were separated according to the emotional resonance method (groups of animals avoiding (altruists) and not avoiding (egotists) the pain cries of partner rats) and neuron activity in the prefrontal areas of the cortex was studied in the right and left hemispheres. Assessments were made of changes in the frequency of nerve cell spike activity (in relation to the baseline activity of neurons in sated animals) in rats subjected to one day of food deprivation and after electrical stimulation of emotionally positive (lateral hypothalamus) and negative (tegmentum of the midbrain) brain structures and after exposure to the pain cries of partner rats. The results of these experiments revealed a series of differences in the cell activities of the two groups of rats. In conditions of hunger, the discharge frequency in the altruists was higher than that in egotists. Cortical neuron responses to positive stimulation were greater than those to negative stimulation in rats of both groups. Intracerebral stimulation produced significantly greater increases in discharge frequency in neurons of both prefrontal areas of the cortex in altruists than in egotists. In both groups of rats, neurons in the right hemisphere responded to emotionally negative stimulation with significantly greater activation than cells in the left hemisphere, while activity in the left hemisphere was greater in conditions of emotionally positive stimulation. Altruists showed significantly greater neuron responses during exposure to pain cries from victim rats in both the right and left hemispheres. The responses of egotists to victim cries were not significantly different from baseline activity levels.     

On the regulation of the expressed L-type calcium channel by cAMP-dependent phosphorylation

The Ca²⁺ channel subunits _1C-a and _1C-b were stably expressed in Chinese hamster ovary (CHO) and human embryonic kidney (HEK) 293 cells. The peak Ba²⁺ current (I _Ba) of these cells was not affected significantly by internal dialysis with 0.1 mM cAMP-dependent protein kinase inhibitor peptide (mPKI), 25 M cAMP-dependent protein kinase catalytic subunit (PKA), or a combination of 25 M PKA and 1 M okadaic acid. The activity of the _1C-b channel subunit expressed stably in HEK 293 cells was depressed by 1 M H 89 and was not increased by superfusion with 5 M forskolin plus 20 M isobutylmethylxanthine (IBMX). The _1C-a·₂·₂/ complex was transiently expressed in HEK 293 cells; it was inhibited by internal dialysis of the cells with 1 M H 89, but was not affected by internal dialysis with mPKI, PKA or microcystin. Internal dialysis of cells expressing the _1C-a·₂·₂/ channel with 10 M PKA did not induce facilitation after a 150-ms prepulse to +50 mV. The Ca²⁺ current (I _Ca) of cardiac myocytes increased threefold during internal dialysis with 5 M PKA or 25 M microcystin and during external superfusion with 0.1 M isoproterenol or 5 M forskolin plus 50 M IBMX. These results indicate that the L-type Ca²⁺ channel expressed is not modulated by cAMP-dependent phosphorylation to the same extent as in native cardiac myocytes.     

Zur Polypeptidketten-Struktur von Immunglobulinen

Zusammenfassung Die Ergebnisse optischer Drehfähigkeitsmessungen an H- und L-Polypeptidketten normaler und Myelomproteine vom G-Typ werden mitgeteilt und mit den optischen Konstanten kompletter G-Proteine verglichen. Durch Kreuzungen zwischen unspezifischen und spezifischen Polypeptidketten werden die Bedingungen einer spezifischen Rekombination geprüft. Sie sind insbesonders dann gegeben, wenn autologe, d.h. vom gleichen Myelomprotein stammende Polypeptidketten gemischt werden. Die Befunde werden in ihrer Beziehung zum Problem der Antikörperspezifität diskutiert.

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Summary The results of measurements of the optical rotatory dispersion on H- and L-polypeptide chains of normal and myeloma proteins are reported Comparisons of the optical parameters with those of the whole G-proteins were performed. By mixing of normal or unspecific and myeloma or specific polypeptide chains the conditions of specific recombination were examined. Specific recombination occurs especially when autologous chains, that means polypeptide chains of an individual myeloma protein, are mixed. The implications of this finding are discussed with respect of the problem of antibody specifity.

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Herrn Prof. Dr.H. E. Bock zum 65. Geburtstag.     

„Leptospira Mini“ ein neuer Serotyp pathogener Leptospiren

Zusammenfassung Auf Grund ihres immunbiologischen Verhaltens gehören die Leptospirenstämme Sari, Ghidorsi und Szwajizak zu demselben serologischen Leptospirentyp, für den der Name Leptospira Mini vorgeschlagen wird.Der Stamm Sari wurde 1942 vonMino sowieVercelli in Italien isoliert. Der Stamm Ghidorsi wurde von uns im Zuge unserer Leptospirenforschungen bei einer Reisfeldarbeiterin der Po-Ebene nachgewiesen. Der Stamm Szwajizak, der vonSmith, Brown, Tonge u. Mitarb. im Jahre 1954 beschrieben wurde, ist in Nord-Qeensland gefunden worden. Der Stamm Sari und Ghidorsi gehören dem kompletten Biotyp (AB), der Stamm Szwajizak dem inkompletten (A) an.Leptospira Mini gehört zur Serogruppe hebdomadis. Ihre Virulenz ist schwach und ihre Bedeutung als Erreger menschlicher Leptospiren-infektionen scheint gering zu sein.     

The role ofγδ T cells in the normal and disordered immune system

Summary A small population of T cells does not express the conventional T cell receptor characterized by the and polypeptide chains (TCR) but instead, two polypeptides termed and (TCR). This alternative receptor is able to recognize antigen. It appears early in T cell ontogeny, but its role in the thymus prior to the availability of TCR remains unclear. In selected sites such as skin or gut TCR predominates in mice which might suggest a role of T cells in the first line of defense against infection, T cells secrete lymphokines and display cytotoxic activity. However, their activation requirements may differ from what is known for T cells since MHC-nonrestricted and also CD4 and CD8 negative T cells have been described. Preferential activation by mycobacterial antigens possibly indicates a special repertoire of the T cells. In various diseases slightly increased numbers of T cells were found, but these preliminary studies have not yet provided evidence for a major pathogenetic role of T cells.List of abbreviations C constant region (immunoglobulin or TCR gene segment) - CD4 cluster of differentiation 4 (mainly on helper cells) - CD8 cluster of differentiation 8 (mainly on cytotoxic cells) - D diversity region (immunoglobulin or TCR gene segment) - DNA desoxyribonucleic acid - IL2 interleukin 2 - J joining region (immunoglobulin or TCR gene segment) - kD kiloDalton - MHC major histocompatibility complex - NK natural killer (cells) - RA rheumatoid arthritis - TCR T cell receptor - V variable region (immunoglobulin or TCR gene segment)     

Evidence for a direct relationship between mitochondrial genome organization and regeneration ability in hexaploid wheat somatic tissue cultures

Summary Embryogenic and non-embryogenic long-term callus cultures of hexaploid wheat exhibit differences in the organization of their mitochondrial genome. Embryogenic and non-embryogenic fractions of callus cultures initiated from immature embryos of the wheat cultivar Chinese Spring have been isolated and subsequently subcultured. DNA-DNA hybridization experiments using labelled cloned wheat mitochondrial DNA fragments have shown that the mitochondrial DNA organization of embryogenic subcultures derived from embryogenic parts of Chinese Spring calli is closely related to that of the initial Chinese Spring calli, while non-embryogenic subcultures derived from non-embryogenic fragments of Chinese Spring calli exhibit a mitochondrial DNA organization similar to that found in non-embryogenic calli derived from cultivar Aquila. In addition, somatic tissue cultures initiated from three other non-embryogenic wheat cultivars (Talent, Thésée and Capitole) display mitochondrial DNA arrangements similar to those found in cultivar Aquila. These results strongly suggest that, in wheat callus cultures, a particular mitochondrial genome organization is correlated with the ability of cultured cells to regenerate whole plants.Abbreviations mtDNA mitochondrial DNA - ctDNA chloroplast DNA - rRNA ribosomal RNA - kb kilobase pair - cv cultivar - 2,4-D 2,4-dichlorophenoxyacetic acid     

The use of electrophoretic karyotypes in the classification of yeasts: Kluyveromyces marxianus and K. lactis

Summary The relationship between varieties marxianus and lactis of Kluyveromyces marxianus was investigated. Strains of these varieties readily form hybrids, but their classification in one species has recently been contested. Enzyme patterns and the GC contents of their DNAs differed significantly. Moreover, DNA-DNA reassociation was less than 15%. Since the generally accepted definition of a species is based on gene exchange, we used two approaches for directly detecting genetic recombination in crosses between representatives of both species. First, multiple marked strains were crossed, and the offspring from the resulting hybrids analyzed. Secondly, the fate of individual chromosomes in identical crosses was followed by comparing the karyotypes of the parent strains, the hybrids formed between them, and the descendants of these hybrids. Karyotypes were obtained by orthogonal-field-altemation gel electrophoresis (OFAGE). Gene exchange was not detected with either method. We therefore concluded that the formation of hybrids, even when they produced viable offspring, was not sufficient to include var. lactis and var. marxianus in one species.     

Hemoglobin heterogeneity under conditions of abnormal erythropoiesis

Experiments on rats showed that changes in the hemoglobin profile during hypoxia are determined by switching of erythropoiesis from the basic to emergency mode. The basic mode of erythropoiesis is typical of the mature organism under normal conditions. The reserve or emergency mode is realized in fetuses, senile animals, and during hypoxia. This mode is characterized by production of large erythrocytes with high content of fetal hemoglobin.     

Anti-5-hydroxytryptamine antibodies: studies on their cross-reactivity in vitro and their immunohistochemical specificity

Summary Antibodies to 5-hydroxytryptamine (5-HT) were obtained from 4 rabbits after injections of 5-HT coupled to bovine serum albumin by means of paraformaldehyde (PF). Two methods were used to monitor the developement of antibodies (AB): the one based on the in vitro competitive binding properties of the antibodies with³(H)5-HT, the other, on their in situ binding properties to endogenous 5-HT, using the peroxidase-anti-peroxidase immunohistochemical technique, applied to paraffin embedded sections of cat brainstem. No pharmacological processing, detergents or proteolytic enzymes were used. The specificity of the antiserum was tested by competitive procedures with 20 analogs using the in vitro and in situ techniques. In vitro studies were performed with 5-HT free analogs and with analogs previously coupled with PF to lysine. Radioimmunological tests showed that the antibodies recognize mainly the ethylamine (CH₂-CH₂-NH₂)-cham of the free analogs and that the best specificity was obtained with the 5-HT conjugate (5-HT-lysine-PF). The results suggest that the hapten is coupled through the phenolic positions C4 or C5. The in situ immunohistochemical extinction assays also revealed a distinct specificity for 5-HT. Possible optical and ultrastructural applications are illustrated in the raphé nuclei of the cat. These results confirm the reliability of radioimmunological tests for studying the specificity of AB directed against haptens, provided that haptens and analogs tested were first chemically transformed to resemble the immunogen (herewith lysine-PF coupling) with regard to its antigenic structure.     

The Involvement of Dopamine in Strengthening Cortical Signals Activating NMDA Receptors in the Striatum (a hypothetical mechanism)

A possible mechanism is proposed for the enhancement/weakening of those cortical signals in the cortex-basal ganglia-thalamus-cortex neural network which induce/do not induce opening of NMDA channels in the spiny neurons of the striatum and which can be regarded as strong/weak in terms of this measure. The mechanism is based on the modulatory influences of dopamine on changes in the efficiency of corticostriatal inputs. In the absence of dopamine, relative increases in the intensity of strong (weak) cortical signals can lead to the induction of long-term potentiation (depression) of corticostriatal synapses. In this case, because of the differently directed influences on thalamic cells of signals passing via strionigral and striopallidal cells, strong signals at the output of the thalamus are weakened, while weak signals are strengthened. Activation of dopamine D₁ (D₂) receptors on strionigral (striopallidal) neurons may facilitate increases in the extent of long-term potentiation/depression (decreases in the extent of long-term potentiation/depression or induction of long-term potentiation/depression). The consequence of this is that strong signals at the output of the thalamus can be strengthened synergistically, while weak signals cab be weakened synergistically. Background cortical signals evoking tonic release of dopamine in the striatum can decrease strengthening because of weakening of the modulatory influence of dopamine on the modification of corticostriatal synapses.     

Sublocalization on chromosome 21 of human interferon-alpha receptor gene and the gene for an interferon-gamma response protein

The cellular responses to alpha and beta interferons (IFN- and -) are mediated through the IFN-/ (type I) receptor, while the response to IFN- is mediated through the IFN- (type II) receptor. The receptors for IFN-/ and IFN- are encoded by genes on human chromosomes 21 and 6q, respectively. The presence of chromosome 21q confers both ligand binding and responsiveness to human IFN-/, whereas chromosome 6q confers binding of Hu-IFN-, but not cellular responsiveness on somatic cell hybrids. Chromosome 6q (i.e., the Hu-IFN- receptor gene) and chromosome 21q are both necessary for the cellular response of somatic cell hybrids (from fibroblasts) to Hu-IFN-. It is conceivable that the factor mediating activity through the IFN- receptor is, in fact, the IFN- receptor, or that the two genes are distinct but part of an interferon response region. Here we more precisely localize on human chromosome 21 the genes for the IFN- receptor and for the factor(s) mediating the action of IFN- through the chromosome 6-encoded receptor. Hamster-human somatic cell hybrids containing various fragments of human chromosome 21 were used. The presence of the human IFN-/ receptor was determined by binding³²P-labeled human IFN- to cells, covalently cross-linking the [³²P]IFN--receptor complex, and analyzing it by SDS-polyacrylamide gel electrophoresis. The presence of the IFN- receptor-related factor mediating cellular responsiveness was determined by HLA induction in hybrid cells containing the IFN- receptor (chromosome 6q), a transfected copy of the human HLA-B7 gene, and various portions of chromosome 21. In all hybrids examined, the two genes cosegregate. Specifically, both genes are localized to the region of chromosome 21 containing the markers D21S58, D21S65, and GART and appear to be proximal to D21S58. The implications for IFN action are discussed.     

Increased levels of α6 integrins are associated with the metastatic phenotype of human breast cancer cells

Integrins play an important role in interactions between cells and the extracellular matrix, and thus have a potential role in metastasis. Expression levels of 6, 1 and 4 integrin sub-units were measured in a panel of human breast cancer cell lines by RT/PCR, immunoprecipitation and flow cytometry. All the lines expressed 6, with the highest levels in the MDA-MB-231 and MDA-MB-435 cells. These grew the most aggressively and were metastastic in nude mice. Low levels of 6 protein were measured in breast cancer cells that were poorly tumorigenic and non-metastatic in nude mice, and there was an inverse relationship between ER and 6 expression. RT/PCR revealed that all lines expressed the 2 isoforms of 6, with the 6A isoform generally more abundant than 6B isoform. Clones of MDA-MB-435 were isolated by sterile sorting for cells with high or low 6 expression, and two variants established from metastases in nude mice were found to differ in 6 expression. When injected into nude mice, the 6-high variants produced significantly more lung metastases than the 6-low variants. 1 was abundant in all lines, while 4 was not detected in MDA-MB-134 cells, and in the MDA-MB-435 cells an alternately spliced variant of 4 was identified. Sequencing of the alternate variant revealed a novel sequence from a splicing event in the cytoplasmic tail of 4. None of the cells with this variant mRNA expressed detectable levels of 4 protein. Our results suggest that high 6 expression in human breast cancer cells is associated with tumorigenicity and metastatic potential.     

A cyclin protein modulates mitosis in the budding yeast Saccharomyces cerevisiae

Summary For the budding yeast Saccharomyces cerevisiae the mitotic cell cycle is coordinated with cell mass at the regulatory step start. The threshold amount of cell mass (reflected as a critical size) necessary for start is proportional to nutrient quality. This relationship leads to a transient accumulation of cells at start, termed nutrient modulation, upon enrichment of nutrient conditions. Nutrient enrichment abruptly increases the critical size needed for start, causing the smaller cells, produced in the previous cell cycle, to be delayed at start while growing larger. Here we show that, in S. cerevisiae, a second cell-cycle step, at mitosis, also exhibits nutrient modulation, and is, therefore, another point of cell-cycle regulation. At both mitosis and start, nutrient modulation was found through mutation to be regulated by the activity of the cyclin-related WHI1 (CLN3) gene product.     

Relation between cell na extrusion and transtubular absorption in the perfused toad kidney: the effect of K,ouabain and ethacrynic acid

Summary Transtubular absorption of Na and Cl, and intracellular ion concentrations were evaluated in toad kidneys perfused with solutions containing K and without K, and in the presence of 1 mM Ouabain and 1 mM Ethacrynic acid. The following values were obtained with 8.5 mM K: Transtubular absorption of Na and Cl68% (percent of filtered load); cell content 294 mole Na, 433 mole K, 100 mole Cl/g solids. Lack of K in the perfusate diminished transtubular absorption to 25% and the cells gain 244 mole Na/g solids, and lose an equimolecular quantity of K. The process is reversible upon raising the K concentration in the perfusate. Ouabain inhibits transtubular absorption to 6%; the cells lose about 110 mole K/g solids, but cellular Na is maintained at the control levels. Ethacrynic acid inhibits transtubular absorption to 3%; the cells approximately double their Na and Cl content, but their K is maintained at the control levels. These observations cannot be explained exclusively in terms of an effect on the distal tubule. Probably proximal as well as distal tubules are involved. A single Na pump seems insufficient to account for all experimental findings. The existence of two separate pumps is therefore proposed.     

Biotin-rich intranuclear inclusions in morule-lacking adenocarcinoma of the gallbladder: a new category of “neoplastic/non-morular” lesions

Biotin-rich intranuclear inclusions, also called optically clear nuclei, are observed in various neoplastic and non-neoplastic lesions, including pregnancy-related endometrium and benign and malignant neoplasms with morular structures. A recent study reported that lesions with biotin-rich intranuclear inclusions can be classified as (non-neoplastic) pregnancy-related endometrial and as (neoplastic) morular category. In the present report, we describe two cases of well-differentiated adenocarcinoma of the gallbladder in which biotin-rich intranuclear inclusions were found without morular structures. Immunohistochemically, as reported previously, the intranuclear inclusions were positive for biotin and two biotin-binding enzymes (pyruvic acid carboxylase and propionyl CoA carboxylase). Intranuclear expression of -catenin was also observed in neoplastic cells with and without intranuclear inclusion. We also detected a frame shift mutation of APC gene in one case but no mutation of -catenin gene in both cases. Although intranuclear expression of -catenin by mutation of APC gene might contribute to carcinogenesis in our cases, the relationships among intranuclear expressions of -catenin, biotin, biotin-binding enzymes and intranuclear inclusions remain unclear. Our cases are the first neoplastic lesions with biotin-rich intranuclear inclusions that lacked morular structures. We propose a new neoplastic/non-morular category for lesions with biotin-rich intranuclear inclusions.